Inhibitors of syk and JAK protein kinases

ABSTRACT

The present invention is directed to compounds of formula I-V and tautomers thereof or pharmaceutically acceptable salts, esters, and prodrugs thereof which are inhibitors of syk kinase. The present invention is also directed to intermediates used in making such compounds, the preparation of such a compound, pharmaceutical compositions containing such a compound, methods of inhibition syk kinase activity, methods of inhibition the platelet aggregation, and methods to prevent or treat a number of conditions mediated at least in part by syk kinase activity, such as undesired thrombosis and Non Hodgkin&#39;s Lymphoma.

CROSS REFERENCES TO RELATED APPLICATIONS

The present application is a continuation of pending U.S. patentapplication Ser. No. 12/386,525 filed Apr. 16, 2009, which applicationclaims the benefit of priority under 35 USC 119(e) of U.S. ProvisionalPatent Application No. 61/120,348, filed Dec. 5, 2008; U.S. ProvisionalPatent Application No. 61/120,346, filed Dec. 5, 2008; U.S. ProvisionalPatent Application No. 61/045,406, filed Apr. 16, 2008; U.S. ProvisionalPatent Application No. 61/045,499, filed Apr. 16, 2008; U.S. ProvisionalPatent Application No. 61/045,399, filed Apr. 16, 2008; and U.S.Provisional Patent Application No. 61/120,341 filed Dec. 5, 2008. Thisapplication is also a continuation of pending U.S. patent applicationSer. No. 13/269,523 filed Oct. 7, 2011, which is a continuation ofpending U.S. patent application Ser. No. 12/386,525 filed Apr. 16, 2009,which application claims the benefit of priority under 35 USC 119(e) ofU.S. Provisional Patent Application No. 61/120,348, filed Dec. 5, 2008;U.S. Provisional Patent Application No. 61/120,346, filed Dec. 5, 2008;U.S. Provisional Patent Application No. 61/045,406, filed Apr. 16, 2008;U.S. Provisional Patent Application No. 61/045,499, filed Apr. 16, 2008;U.S. Provisional Patent Application No. 61/045,399, filed Apr. 16, 2008;and U.S. Provisional Patent Application No. 61/120,341 filed Dec. 5,2008. The disclosures of each of the foregoing applications are herebyincorporated herein by reference.

BACKGROUND OF THE INVENTION

This invention is directed to pyrimidine-5-carboxamide compounds whichact as inhibitors of Spleen tyrosine kinase (syk) and/or 'JAK kinases.This invention is also directed to pharmaceutical compositionscontaining the pyrimidine-5-carboxamide compounds and methods of usingthe compounds or compositions to treat a condition characterized byother indications. The invention is also directed to methods of makingthe compounds described herein.

STATE OF THE ART

Protein kinases constitute a large family of structurally relatedenzymes that are responsible for the control of a variety of signaltransduction processes within cells (see, e.g., Hardie and Hanks, TheProtein Kinase Facts Book, I and II, Academic Press, San Diego, Calif.,1995). Protein kinases are thought to have evolved from a commonancestral gene due to the conservation of their structure and catalyticfunction. Almost all kinases contain a similar 250-300 amino acidcatalytic domain. The kinases can be categorized into families by thesubstrates they phosphorylate (e.g., protein-tyrosine,protein-serine/threonine, lipids, etc.). Sequence motifs have beenidentified that generally correspond to each of these families (see,e.g., Hanks & Hunter, (1995), FASEB J. 9:576-596; Knighton et al.,(1991), Science 253:407-414; Hiles et al., (1992), Cell 70:419-429; Kunzet al., (1993), Cell 73:585-596; Garcia-Bustos et al., (1994), EMBO J.13:2352-2361).

Many diseases are associated with abnormal cellular responses triggeredby protein kinase-mediated events. These diseases include autoimmunediseases, inflammatory diseases, bone diseases, metabolic diseases,neurological and neurodegenerative diseases, cancer, cardiovasculardiseases, allergies, asthma, alzheimer's disease and hormone-relateddiseases. As a consequence, there has been substantial efforts inmedicinal chemistry to find inhibitors of protein kinases for use astherapeutic agents.

Immunoreceptor tyrosine activation motif (ITAM)-mediated signaling hasemerged as a primary event in signaling pathways responsible for humanpathologies. ITAM-mediated signaling is responsible for relayingactivation signals initiated at classical immune receptors such asT-cell receptors, B-cell receptors, Fc receptors in immune cells and atGPVI and FcγRIIa in platelets to downstream intracellular molecules suchas syk and ZAP-70 (Underhill, D. M and Goodridge, H. S., TrendsImmunol., 28:66-73, 2007).

The binding of a ligand to an ITAM-containing receptor triggerssignaling events which allows for the recruitment of proteins from afamily of nonreceptor tyrosine kinases called the Src family. Thesekinases phosphorylate tyrosine residues within the ITAM sequence, aregion with which the tandem SH2 domains on either syk or ZAP-70interact.

Syk, along with Zap-70, is a member of the syk family of proteintyrosine kinases. The interaction of syk or ZAP-70 with diphosphorylatedITAM sequences induces a conformation change in the kinases that allowsfor tyrosine phosphorylation of the kinase itself. Phosphorylated Sykfamily members activate a multitude of downstream signaling pathwayproteins which include Src homology 2 (SH2) domain containingleukocyte-specific phosphoprotein of 76 kDa (SLP-76), Linker ofActivation of T-cells (LAT) and PLC (phospholipase C)γ2.

Human pathologies attributed to dysfunctional ITAM-mediated signalinginclude autoimmune diseases such as rheumatoid arthritis, systemiclupus, multiple sclerosis, hemolytic anemia, immune-thrombocytopeniapurpura, and heparin-induced thrombocytopenia and arteriosclerosis.Interestingly, many of the above mentioned diseases are thought to occurthrough crosslinking of Fc receptors by antibodies which, via syk,activate a signaling cascade in mast, basophil and other immune cellsthat result in the release of cell mediators responsible forinflammatory reactions. The release of mediators and the production ofcytokines in IgE stimulation-dependent allergic and inflammatoryreactions from mast cells and basophiles can be controlled by inhibitingthe tyrosine kinase activity of syk (Rossi, A. B. et al., J Allergy ClinImmunol., 118:749-755, 2006). In immune-thrombocytopenia, antibody boundplatelets are cleared by the spleen by an Fc receptor/ITAM/syk-mediatedprocess (Crow, A. R. et al., Blood, 106:abstract 2165, 2005).Drug-induced thrombocytopenia, caused by heparin-platelet factor 4immune complexes that activate platelet FcγRIIa, also involve syksignaling downstream of receptor engagement (Reilly, M. P., Blood,98:2442-2447, 2001).

Platelet agonists induce inside-out integrin signaling resulting infibrinogen binding and platelet aggregation. This initiates outside-insignaling which produces further stimulation of platelets. syk isactivated during both phases of integrin signaling, and inhibition ofsyk is shown to inhibit platelet adhesion to immobilized proteins (Law,D. A. et al., Blood, 93:2645-2652, 1999). Release of arachidonic acidand serotonin and platelet aggregation induced by collagen are markedlyinhibited in platelets derived from syk deficient mouse (Poole, A. etal., EMBO J., 16:2333-2341, 1997). Thus syk inhibitors may also possessanticoagulation action.

Because of the role syk plays in Ig-induced platelet activations, it islikely to be important in arteriosclerosis and restenosis.Arteriosclerosis is a class of diseases characterized by the thickeningand hardening of the arterial walls of blood vessels. Although all bloodvessels are susceptible to this serious degenerative condition, theaorta and the coronary arteries serving the heart are most oftenaffected. Arteriosclerosis is of profound clinical importance since itcan increase the risk of heart attacks, myocardial infarctions, strokes,and aneurysms.

The traditional treatment for arteriosclerosis includes vascularrecanalization procedures for less-serious blockages and coronary bypasssurgery for major blockages. A serious shortcoming of intravascularprocedures is that, in a significant number of treated individuals, someor all of the treated vessels restenose (i.e., re-narrow). For example,restenosis of an atherosclerotic coronary artery after PTCA occurs in10-50% of patients undergoing this procedure and subsequently requireseither further angioplasty or a coronary artery bypass graft.Furthermore, restenosis of an atherosclerotic coronary artery afterstenting occurs in 10-20% of patients undergoing this procedure andsubsequently requires repeat treatments to maintain adequate blood flowthrough the affected artery. Restenosis generally occurs in a relativelybrief time period, e.g., roughly less than six months, after treatment.

While the exact hormonal and cellular processes promoting restenosishave not been determined, restenosis is thought to be due in part tomechanical injury to the walls of the blood vessels caused by theballoon catheter or other intravascular device. For example, the processof PTCA, in addition to opening the obstructed artery, also injuresresident coronary arterial smooth muscle cells (SMCs). In response tothis injury, adhering platelets, infiltrating macrophages, leukocytes,or the smooth muscle cells themselves release cell-derived growthfactors such as platelet-derived growth factor (PDGF), with subsequentproliferation and migration of medial SMCs through the internal elasticlamina to the area of the vessel intima. Further proliferation andhyperplasia of intimal SMCs and, most significantly, production of largeamounts of extracellular matrix over a period of three to six monthsresults in the filling in and narrowing of the vascular space sufficientto significantly obstruct blood flow.

In addition to the role syk plays in Ig-induced platelet activations,syk plays a very important role in collagen-mediated signaling. Theprimary adhesive protein responsible for platelet adhesion andactivation is collagen. Collagen is a filamentous protein containedwithin the fibrotic caps of atheromas which becomes exposed to bloodduring plaque rupture. Collagen functions initially by binding vonWillebrand factor which tethers platelets through binding plateletmembrane GPIb. Collagen functions secondarily by engaging the twocollagen receptors on platelets, GPVI and integrin α2β1.

GPVI exists in platelet membranes as a complex with FcRγ, an interactionrequired for the expression of GPVI. Activation of FcγRIIa on plateletsresults in platelet shape change, secretion and thrombosis. Signaling bythe GPVI/FcRγ complex is initiated by tyrosine phosphorylation of theITAM domain of FCRγ followed by the recruitment of syk.

Activation of GPVI leads to induction of multiple platelet functionsincluding: activation of integrins α2β1 to achieve firm plateletadhesion, and GP IIb-IIIa which mediates platelet aggregation andthrombosis growth; platelet secretion, allowing for the delivery ofinflammatory proteins such as CD40L, RANTES and TGFβ to the vessel wall;and the expression of P-selectin which allows for the recruitment ofleukocytes. Therefore, it is believed that syk inhibitors can inhibitthrombotic events mediated by platelet adhesion, activation andaggregation.

It has been reported that the tyrosine phosphorylation of intracellularprotein (activation) induced by stimulation of a receptor for IgGantibody, FcγR, and the phagocytosis mediated by FcγR are considerablyinhibited in macrophages derived from syk deficient mouse (Crowley, M.T. et al., J. Exp. Med., 186:1027-1039, 1997). This suggests that sykhas a markedly important role in the FcγR-mediated phagocytosis ofmacrophages.

It has also been reported that an antisense oligonucleotide of syksuppresses the apoptosis inhibition of eosinophils induced by GM-CSF(Yousefi, S. et al., J. E. Med., 183:1407-1414, 1996), showing that sykis essential for the life extending signal of eosinophils caused byGM-CSF and the like. Since life extension of eosinophils is closelyrelated to the transition of diseases into a chronic state in allergicdisorders, such as asthma, syk inhibitors can also serve as therapeuticagents for chronic eosinophilic inflammation.

Syk is important for the activation of B-cells via a B-cell antigenreceptor and is involved in the phosphatidylinositol metabolism andincrease in the intracellular calcium concentration caused by theantigen receptor stimulation (Hutchcroft, J E. et al., J. Biol. Chem.,267:8613-8619, 1992; and Takata, M. et al., EMBO J., 13:1341-1349,1994). Thus, syk inhibitors may be used to control the function ofB-cells and are, therefore, expected to serve as therapeutic agents forantibody-related diseases.

Syk binds to a T-cell antigen receptor, quickly undergoes tyrosinephosphorylation through crosslinking of the receptor and synergisticallyacts upon intracellular signals mediated by Src tyrosine kinases such asLck (Couture, C. et al., Proc. Natl. Acad. Sci. USA, 91:5301-5305, 1994;and Couture, C. et al., Mol. Cell. Biol., 14:5249-5258, 1994). syk ispresent in mature T-cell populations, such as intraepithelial γδ T-cellsand naïve αβ T-cells, and has been reported to be capable ofphosphorylation of multiple components of the TCR signaling cascade(Latour, S. et. al., Mol Cell Biol., 17:4434-4441, 1997). As aconsequence, syk inhibitors may serve as agents for inhibiting cellularimmunity mediated by T-cell antigen receptor.

Recent comparative genomic hybridization studies have identified syk asanother gene important in the pathogenesis of Mantle Cell Lymphoma (MCL)(Chen, R. et al. Journal of Clinical Oncology, 2007 ASCO Annual MeetingProceedings (Post-Meeting Edition). Vol 25, No 18S (June 20 Supplement),2007: 8056). MCL represents 5-10% of all non-Hodgkins lymphomas and itis a difficult form of lymphoma to treat. It has the worst prognosisamong the B cell lymphomas with median survival of three years. It hasbeen reported that Syk is overexpressed in MCL (Rinaldi, A, et. al, Br.J. Haematol., 2006; 132:303-316) and that Syk mediates mTOR (mammaliantarget of Rapamycin) survival signals in follicular, mantel cell,Burkitt's, and diffuse large B-cell non-Hodgkin's lymphomas (Leseux, L.,et. al, Blood, 2006; 108:4156-4162).

Several lines of evidence suggest that many B-cell lymphomas depend uponB-cell receptor (BCR)-mediated survival signals. BCR signaling inducesreceptor oligomerization and phosphorylation of Igα and β immunoreceptortyrosine-based activated motifs by SRC family kinases. ITAMphosphorylation results in the recruitment and activation of syk thatinitiates downstream events and amplifies the original BCR signal. Giventhe role of tonic BCR signaling in normal B cell and syk-dependentsurvival of non-Hodgkins lymphoma cell lines in vitro (Chen, L., et. al,Blood, 2006; 108:3428-3433), syk inhibition is a promising rationaltreatment target for certain B-cell lymphomas and chronic lymphocyticleukemia (CLL) (Stefania Gobessi, Luca Laurenti, Pablo Longo, LauraCarsetti, Giuseppe Leone, Dimitar G. Efremov, Constitutive activation ofthe protein tyrosine kinase Syk in Chronic Lymphocytic Leukemia B-cells,Blood, 2007, 110, Abstract 1123). Recent data shows that administrationof a multikinase inhibitor which inhibits syk, may have significantclinical activity in CLL patients (Friedberg J W et al, Blood 2008;112(11), Abstract 3).

The oncogenic potential of the spleen tyrosine kinase (Syk) has beendescribed in a number of different settings. Clinically, Sykover-expression is reported in Mantle Cell Lymphoma (Rinaldi, A, et. al,Br. J. Haematol., 2006; 132:303-316) and the TEL-Syk fusion protein(Translocated ETS Leukemia) generated by a chromosomal translocation(t(9;12)(q22;p12)) leads to increased Syk activity and is associatedwith myelodysplastic syndrome (Kuno, Y., et. al, Blood, 2001;97:1050-1055). Leukemia is induced in mice by adoptively transferringbone marrow cells that express human TEL-Syk (Wossning, T., JEM, 2006;203:2829-2840). Further, in mouse primary bone marrow cells,over-expression of Syk results in IL-7 independent growth in culture(Wossning, T., et. al, JEM, 2006; 203:2829-2840).

Interestingly, Syk signaling appears to be required for B-celldevelopment and survival in humans and mouse. Inducible loss of theB-cell receptor (Lam, K., et. al, Cell, 1997; 90:1073-1083) or Igα(Kraus, M., et. al, Cell, 2004; 117:787-800) results in loss ofperipheral B-cells in mice. Over-expression of the protein tyrosinephosphatase PTP-RO, which is known to negatively regulate Syk activity,inhibits proliferation and induces apoptosis in cell lines derived fromnon-Hodgkin's lymphomas (Chen, L., et. al, Blood, 2006; 108:3428-3433).Finally, B-cell lymphomas rarely exhibit loss of BCR expression, andanti-idiotype therapy rarely leads to resistance (Kuppers, R. Nat RevCancer, 2005; 5:251-262).

Engagement of the antigen-specific B cell receptor (BCR) activatesmultiple signaling pathways that ultimately regulate the cellsactivation status, promoting survival and clonal expansion. Signalingthrough the BCR is made possible by its association with two othermembers of the immunoglobulin super-family; Igα and Igβ, each bearing animmuno-tyrosine based activation motif (ITAM) (Jumaa, Hendriks et al.Annu Rev Immunol 23: 415-45 (2005). The ITAM domain is directlyphosphorylated by Src family kinases in response to BCR engagement. Thespleen tyrosine kinase (Syk) docks with and phosphorylates the ITAM, aprocess that enhances its kinase activity, resulting in Sykautophosphorylation and tyrosine phosphorylation of multiple downstreamsubstrates (Rolli, Gallwitz et al. Mol Cell 10(5): 1057-69 (2002). Thissignaling pathway is active in B cells beginning at the transition frompro- to pre-B cell stage of development, when the newly formed pre-BCRis expressed. In fact, B cell development arrests at the pro-B cellstage in Syk knockout mice (Cheng, Rowley et al. 1995; Turner, Mee etal. Nature 378(6554): 303-6 (1995). Inducible loss of the B cellreceptor (Lam, Kuhn et al. Cell 90(6): 1073-83 (1997) or Igα (Kraus,Alimzhanov et al. Cell 117(6): 787-800 (2004) results in loss ofperipheral B cells in mice. Human B cells also appear to require Syk forproliferation and survival. Over-expression of the protein tyrosinephosphatase PTP-RO, a negative regulator of Syk activity, inhibitsproliferation and induces apoptosis in cell lines derived fromnon-Hodgkin's lymphomas (NHL) (Chen, Juszczynski et al. Blood 108(10):3428-33 (2006). Knock down of Syk by siRNA in the NHL line SUDHL-4 ledto a block in the G1/S transition of the cell cycle (Gururajan, Dasu etal. J Immunol 178(1): 111-21 (2007). Together, these data suggest thatSyk signaling is required for the development, proliferation, and evensurvival of human and mouse B cells.

Conversely, the oncogenic potential of Syk has been described in anumber of different settings. Clinically, Syk over-expression isreported in Mantle Cell Lymphoma (Rinaldi, Kwee et al. Br J Haematol132(3): 303-16 (2006) and the TEL-Syk fusion protein (Translocated ETSLeukemia) generated by a chromosomal translocation (t(9;12)(q22;p12))leads to increased Syk activity and is associated with myelodysplasticsyndrome (Kuno, Abe et al. Blood 97(4): 1050-5 (2001). Leukemia isinduced in mice by the adoptive transfer of bone marrow cells thatexpress human TEL-Syk (Wossning, Herzog et al. J Exp Med 203(13):2829-40 (2006). Further, in mouse primary bone marrow cells,over-expression of Syk results in IL-7 independent growth in culture(Wossning, Herzog et al. 2006). Consistently, Syk was reported tomediate mTOR (mammalian target of Rapamycin) survival signals infollicular, mantle cell, Burkitt's, and diffuse large B-cell NHL(Leseux, Hamdi et al. Blood 108(13): 4156-62 (2006). Additional recentstudies also suggest that Syk-dependant survival signals may play a rolein B-cell malignancies, including DLBCL, mantle cell lymphoma andfollicular lymphoma (Gururajan, Jennings et al. 2006; Irish, Czerwinskiet al. J Immunol 176(10): 5715-9 (2006). Given the role of tonic BCRsignaling in normal B cells and Syk-dependent survival of NHL cell linesin vitro, the specific inhibition of Syk may prove promising for thetreatment of certain B-cell lymphomas.

Recently, R406 (Rigel Pharmaceuticals) was reported to inhibit ITAMsignaling in response to various stimuli, including FcεR1 and BCRinduced Syk activation (Braselmann, Taylor et al. J Pharmacol Exp Ther319(3): 998-1008 (2006). Interestingly, this ATP-competitive inhibitorof Syk was also active against Flt3, cKit, and JAK kinases, but notagainst Src kinsase (Braselmann, Taylor et al. 2006). Activatingmutations to Flt3 are associated with AML and inhibition of this kinaseis currently under clinical development (Burnett and Knapper HematologyAm Soc Hematol Educ Program 2007: 429-34 (2007). Over-activation of thetyrosine kinase cKit is also associated with hematologic malignancies,and a target for cancer therapy (Heinrich, Griffith et al. Blood 96(3):925-32 (2000). Similarly, JAK3 signaling is implicated in leukemias andlymphomas, and is currently exploited as a potential therapeutic target(Heinrich, Griffith et al. 2000). Importantly, the multi-kinaseinhibitory activity of R406 attenuates BCR signaling in lymphoma celllines and primary human lymphoma samples, resulting in apoptosis of theformer (Chen, Monti et al. Blood 111(4): 2230-7 (2008). Further, a phaseII clinical trial reported favorable results by this compound inrefractory NHL and chronic lymphocytic leukemia (Friedberg J W et al,Blood 2008; 112(11), Abstract 3). Although the precise mechanism ofaction is unclear for R406, the data suggest that inhibition of kinasesthat mediate survival signaling in lymphocytes is clinically beneficial.

Additional recent studies also suggest that syk-dependant survivalsignals may play a role in B-cell malignancies, including DLBCL, mantlecell lymphoma and follicular lymphoma (see e.g., S. Linfengshen et al.Blood, February 2008; 111: 2230-2237; J. M. Irish et al. Blood, 2006;108: 3135-3142; A. Renaldi et al. Brit J. Haematology, 2006; 132:303-316; M. Guruoajan et al. J. Immunol, 2006; 176: 5715-5719; L. Laseuxet al. Blood, 2006; 108: 4156-4162.

Patents and patent applications describing substituted pyrimidinediaminecompounds include: U.S. application Ser. No. 10/355,543 filed Jan. 31,2003 (US2004/0029902A1), international application Serial No.PCT/US03/03022 filed Jan. 31, 2003 (WO 03/063794), U.S. application Ser.No. 10/631,029 filed Jul. 29, 2003, international application Serial No.PCT/US03/24087 (WO 04/014382), U.S. application Ser. No. 10/903,263filed Jul. 30, 2004, and international application Serial No.PCT/US2004/24716 (WO 05/016893), the disclosures of which areincorporated herein by reference. Substituted pyrimidinediaminecompounds are also described in international patent applicationpublication numbers: WO 02/059110, WO 03/074515, WO 03/106416, WO03/066601, WO 03/063794, WO 04/046118, WO 05/016894, WO 05/122294, WO05/066156, WO 03/002542, WO 03/030909, WO 00/39101, WO 05/037800 andU.S. Pat. Pub. No. 2003/0149064.

While progress has been made in this field, there remains a need in theart for compounds that inhibit syk kinase, as well as for methods fortreating conditions in a patient, such as restenosis, thrombosis, and/orinflammation that can benefit from such inhibition. Moreover, theavailability of compounds that selectively inhibit one of these kinasesas compared to other kinases would also be desirable. The presentinvention satisfies this and other needs.

BRIEF SUMMARY OF THE INVENTION

The present invention provides novel compounds having activity asinhibitors of syk activity (also referred to herein as “syk inhibitors”)kinase activity (also referred to herein as “JAK inhibitors”), as wellas to methods for their preparation and use, and to pharmaceuticalcompositions containing the same. Such compounds have the followingstructure (I):

or a pharmaceutically acceptable salt thereof, wherein D¹, E¹ and Y¹ areas defined below.

The present invention also provides a pharmaceutical compositioncomprising a therapeutically effective amount of a compound of formulaI, or a pharmaceutical acceptable salt thereof, and a pharmaceuticallyacceptable carrier and/or diluent.

The compounds of the present invention have utility over a wide range oftherapeutic applications, and may be used to treat a variety ofconditions, mediated at least in part by syk activity, in both men andwomen, as well as a mammal in general (also referred to herein as a“subject”). For example, such conditions include, but are not limitedto, those associated with cardiovascular disease, inflammatory diseaseor autoimmune disease. More specifically, the compounds of the presentinvention have utility for treating conditions or disorders including,but not limited to: restenosis, thrombosis, inflammation, heparininduced thrombocytopenia, dilated cardiomyopathy, sickle cell disease,atherosclerosis, myocardial infarction, vascular inflammation, unstableangina, acute coronary syndromes, allergy, asthma, rheumatoid arthritis,B-cell mediated diseases such as Non Hodgkin's lymphoma,anti-phospholipid syndrome, lupus, psoriasis, multiple sclerosis, endstage renal disease, hemolytic anemia, immune thrombocytopenic purpura,and chronic lymphocytic leukemia. Thus, in one embodiment, methods aredisclosed which include the administration of an effective amount of acompound of formula (I), typically in the form of a pharmaceuticalcomposition, to a subject in need thereof.

The conditions associated with cardiovascular disease is selected fromthe group consisting of acute coronary syndrome, myocardial infarction,unstable angina, refractory angina, occlusive coronary thrombosisoccurring post-thrombolytic therapy or post-coronary angioplasty, athrombotically mediated cerebrovascular syndrome, embolic stroke,thrombotic stroke, transient ischemic attacks, venous thrombosis, deepvenous thrombosis, pulmonary embolism, coagulopathy, disseminatedintravascular coagulation, thrombotic thrombocytopenic purpura,thromboangiitis obliterans, thrombotic disease associated withheparin-induced thrombocytopenia, thrombotic complications associatedwith extracorporeal circulation, thrombotic complications associatedwith instrumentation such as cardiac or other intravascularcatheterization, intra-aortic balloon pump, coronary stent or cardiacvalve, and conditions requiring the fitting of prosthetic devices.

The present invention also provides a method for inhibiting the sykkinase activity of a blood sample comprising contacting said sample witha compound of the present invention.

The present invention further provides compounds in purified forms, aswell as chemical intermediates.

These and other aspects, objects, features and advantages of theinvention will be apparent upon reference to the following detaileddescription and figures. To this end, various references are set forthherein which describe in more detail certain background information,procedures, compounds and/or compositions, and are each herebyincorporated by reference in their entirety.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows how Syk serves as a key mediator of Fc receptor mediatedsignaling in cellular biology and multiple diseases.

FIG. 2 shows how gene targeting of Syk indicated that Syk serves as akey mediator in arterial platelet biology and a selective target fortreating arterial thrombosis.

FIG. 3 shows a general synthesis of compounds of the present invention.

FIGS. 4A, B and C provides tables 2A, B and C illustrating compounds ofthe present invention and syk IC₅₀s.

FIG. 5 provides table 3 illustrating compounds of the present inventionand syk IC₅₀s.

FIG. 6 provides table 4 illustrating compounds of the present inventionand syk IC₅₀s.

FIGS. 7A and 7B provide table 5 A and B illustrating compounds of thepresent invention and syk IC₅₀s.

FIG. 8 shows a series of compounds that were identified to selectivelyinhibit Syk in purified kinase assays. FIG. 8A) Compounds (example 596and example 87 and P420-89) were screened at 300 nM against theMillipore purified kinase panel (270 kinases tested with 10 μM ATP) todetermine potency and selectivity for Syk. Data are represented as aheat-map, defined at the bottom. FIG. 8B) Subset of the purified kinasesthat had >80% inhibition by any of the three compounds. example 596 onlyinhibited Syk and MLK1. example 87 at 50 nM (˜10× greater than its SykIC50) only inhibited Syk. P420-89 inhibited multiple kinases, includingSyk, JAK2 and JAK3. The IC50 of Syk inhibition is reported for eachcompound on the left of the heat map. Percent kinase inhibition is givenin each panel within the heat-map.

FIG. 9 shows the selective inhibition of Syk in non-Hodgkin's Lymphomacell lines. B cells were stimulated with anti-BCR antibody in thepresence of the indicated concentrations of Syk specific inhibitorsexample 596 and example 87 (FIG. 9A and FIG. 9B) or the dual Syk/JAKinhibitor P420-89 (FIG. 9C). Western blot analyses of whole cell lysateswere then performed to evaluate Syk kinase activity (pBLNK Y84 and totalBLNK; top two gels) and Src kinase activity (pSyk Y352 and total Syk;bottom two gels). Experiments were performed 2-3 times each, bar graphsrepresent mean±S.D. of pBLNK Y84. The calculated IC50s of Syk kinaseinhibition are presented above the graphs.

FIG. 10 provides a comparison of Syk-Specific and Dual Syk/JAKInhibition in NHL Cell Lines. B cells were stimulated with anti-BCR(FIG. 10A), or IL-4 (FIG. 10B) for 15 min in the presence of variousconcentrations of each inhibitor, as indicated. Cells were thenevaluated for inhibition of signaling pathways by phospho-flowcytometry. FIG. 10A) bar graphs (mean±S.D., n=3) representing Srcactivity (pSyk Y352 MFI) and Syk activity (pERK Y204 MFI) following BCRstimulation under the various treatment conditions. FIG. 10B) Bar graphsdepicting pSTAT-6 Y641 MFI (mean±S.D., n=3) following stimulation withIL-4 in the presence of various concentrations of each inhibitor, asindicated.

FIG. 11 shows how Syk-specific inhibitors disrupt proliferation andsurvival of and induces apoptosis in NHL cell lines. The Syk-dependent“BCR type” and Syk-independent “non-BCR type” NHL cell lines werepreviously described (Polo, Juszczynski et al. Proc Natl Acad Sci USA104(9): 3207-12 (2007). FIG. 11A) Cells were treated for 72 h with 1 and3 μM of the Syk-specific inhibitor example 87. Apoptosis was determinedby FACS analysis of active caspase 3; data is represented as histograms.FIG. 11B) Additional cell lines were tested for sensitivity to Sykspecific (example 596 and example 87) versus dual Syk/JAK (P420-89)inhibition. Bar graphs represent mean±S.D. (n=3) of the percent ofcaspase 3 positive cells following each condition.

FIG. 12A shows how selective inhibition of Syk prevents BCR-inducedactivation of mouse primary B cells. FIG. 12B provides a comparison ofSyk-Specific and Dual Syk/JAK Inhibition in NHL Cell Lines. B cells werestimulated with anti-BCR (FIG. 12B) for 15 min in the presence ofvarious concentrations of each inhibitor, as indicated. Cells were thenevaluated for inhibition of signaling pathways by phospho-flowcytometry. FIG. 12C) Flow cytometry plots (mean±S.D., n=3) representingSrc activity (pSyk Y352 MFI) and Syk activity (pERK Y204 MFI) followingBCR stimulation under the various treatment conditions.

FIG. 13 shows the efficacy in a mouse arthritis model by specificinhibition of Syk

FIG. 14 shows how the histopathology in a mouse model confirms theclinical score for a Syk specific inhibitor.

FIG. 15 shows the dose dependent effect of inhibition of Syk/JAK in aMouse Arthritis Model:

FIG. 16 provides a mouse immune thrombocytopenia model

FIG. 17 shows how Syk Inhibition prevents NHL tumor formation in axenograft mouse model with a NHL cell line. Selective Syk inhibitorexample 87 prevents tumor formation. Syk inhibition prevents NHL tumorformation in xenograft mice. Mice were dosed twice daily with 10, 15, or20 mg/kg example 87 or vehicle control beginning the day of tumor cellinoculation. A) Tumor weights were determined at 4 weekspost-inoculation for each treatment condition. Statistical differencesrelative to vehicle control are depicted as P values within the graph.B) Bar graphs depict normal blood cell counts (mean±S.D., n=13 to 15) ofmice treated with each concentration of example 87 or vehicle control.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the below terms have the following meanings unlessspecified otherwise:

1. Abbreviations and Definitions

The abbreviations used herein are conventional, unless otherwisedefined. The following abbreviations are used: AcOH=acetic acid,AIBN=azobisisobutyronitrile (also azobisisobutylonitrile), aq.=aqueous,Boc=t-butylcarboxy, Bz—benzyl,BOP=benzotriazol-1-yloxytris(dimethylamino)-phosphoniumhexafluorophosphate, BPO=benzoyl peroxide, nBuOH=n-butanol,CBr₄=tetrabromomethane, mCPBA=m-chloroperoxybenzoic acid, CH₂Cl₂ orDCM=dichloromethane, Cs₂CO₃=cesium carbonate, CuCl₂=copper chloride;DPPA=diphenyl phosphoryl azide; DIBAL=diisobutylaluminum hydride,DIEA=Hunig's base or diisopropyl ethylamine, DME=dimethyl ether,DMF=dimethyl formamide, DMSO=dimethyl sulfoxide, Et₃N=triethylamine,EtOAc=ethyl acetate, g=gram, HATU=2-(1H7-Azabenzotriazol-1-yl)-1,1,3,3-tetramethyl uronium hexafluorophosphate,H₂=hydrogen; H₂O=water; HBr=hydrogen bromide; HCl=hydrogen chloride,HIV=human immunodeficiency virus, HPLC=high pressure liquidchromatography, h=hour, IgE=immunoglobulin E, IC₅₀=The concentration ofan inhibitor that is required for 50% inhibition of an enzyme in vitro,IPA=isopropyl alcohol, kg=kilogram, KCN=potassium cyanide, KOH=potassiumhydroxide, K₂PO₄=potassium phosphate, LDA=lithium diisopropylamine,LiAlH₄=lithium aluminum hydride=LiOH: lithium hydroxide;MeCN=acetonitrile; MS=Mass Spec, m/z=mass to charge ratio, MHz=MegaHertz, MeOH=methanol, μM=micromolar, μL=microliter, mg=milligram,mm=millimeter, mM=millimolar, mmol=millimole, mL=milliliter,mOD/min=millioptical density units per minute, min=minute, M=molar,Na₂CO₃=sodium carbonate, ng=nanogram, NaHCO₃=sodium bicarbonate;NaNO₂=sodium nitrite; NaOH=sodium hydroxide; Na₂S₂O₃=sodium bisulfate;Na₂SO₄=sodium sulfate; NBS=N-bromosuccinamide; NH₄Cl=ammonium chloride;NH₄OAc=ammonium acetate; NaSMe=sodium methylthiolate,NBS=N-bromosuccinamide, n-BuLi=n-butyl lithium, nm=nanometer,nM=nanomolar, N=Normal, NMP=N-methylpyrrolidine, NMR=nuclear magneticresonance, Pd/C=palladium on carbon,Pd(PPh₃)₄=Tetrakis-(triphenyl-phosphine)-palladium, pM=picomolar,Pin=pinacolato, PEG=polyethylene glycol, PPh₃ or Ph₃P=triphenylphosphine, RLV=Raucher leukemia virus, Ra—Ni=Rainey Nickel,SOCl₂=thionyl chloride, RT=room temperature, TEA=triethylamine,THF=tetrahydrofuran, TFA=trifluoroacetic acid, TLC=thin layerchromatography, TMS=trimethylsilyl, Tf=trifluoromethylsulfonyl andTSC=trisodium citrate.

It is noted here that as used in this specification and the appendedclaims, the singular forms “a,” “an,” and “the” include plural referenceunless the context clearly dictates otherwise.

“Alkyl,” by itself or as part of another substituent, means, unlessotherwise stated, a straight or branched chain, fully saturatedaliphatic hydrocarbon radical having the number of carbon atomsdesignated. For example, “C₁₋₈alkyl” refers to a hydrocarbon radicalstraight or branched, containing from 1 to 8 carbon atoms that isderived by the removal of one hydrogen atom from a single carbon atom ofa parent alkane. The phrase “unsubstituted alkyl” refers to alkyl groupsthat do not contain groups other than fully saturated aliphatichydrocarbon radicals. Thus the phrase includes straight chain alkylgroups such as methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl,octyl, nonyl, decyl, undecyl, dodecyl and the like. The phrase alsoincludes branched chain isomers of straight chain alkyl groups such asisopropyl, t-butyl, isobutyl, sec-butyl, and the like. Representativealkyl groups include straight and branched chain alkyl groups having 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. Furtherrepresentative alkyl groups include straight and branched chain alkylgroups having 1, 2, 3, 4, 5, 6, 7 or 8 carbon atoms.

“Alkenyl” by itself or as part of another substituent refers to astraight or branched chain, which may be mono- or polyunsaturated,having the number of carbon atoms designated. For example, “C₂-C₈alkenyl” means an alkenyl radical having from 2, 3, 4, 5, 6, 7 or 8atoms that is derived by the removal of one hydrogen atom from a singlecarbon atom of a parent alkane. Examples include, but are not limited tovinyl, 2-propenyl i.e. —CH═C(H)(CH₃), —CH═C(CH₃)₂, —C(CH₃)═C(H)₂,—C(CH₃)═C(H)(CH₃), —C(CH₂CH₃)═CH₂, butadienyl e.g. 2-(butadienyl),pentadienyl e.g. 2,4-pentadienyl and 3-(1,4-pentadienyl), andhexadienyl, among others, and higher homologs and stereoisomers thereof.A “substituted” alkenyl group includes alkenyl groups in which anon-carbon or non-hydrogen atom is bonded to a carbon double bonded toanother carbon and those in which one of the non-carbon or non-hydrogenatoms is bonded to a carbon not involved in a double bond to anothercarbon. Each site of unsaturation may be either cis or transconfiguration about the double bond(s).

The term “alkynyl”, by itself or as part of another substituent, means astraight or branched chain hydrocarbon radical, which may be mono- orpolyunsaturated, having the number of carbon atoms designated. Forexample, “C₂-C₈ alkynyl” means an alkynyl radical having from 2 to 8carbon atoms that is derived by the removal of one hydrogen atom from asingle carbon atom of a parent alkane. “Unsubstituted alkynyl” refers tostraight and branched chain groups such as those described with respectto unsubstituted alkyl groups as defined above, except that at least onetriple bond exists between two carbon atoms. Examples include, but arenot limited to ethynyl e.g. —Ca≡C(H), 1-propynyl e.g. —C≡C(CH₃),—C≡C(CH₂CH₃), —C(H₂)C≡C(H), —C(H)₂C≡C(CH₃), and —C(H)₂C≡C(CH₂CH₃) amongothers, and higher homologs and isomers thereof. A “substituted” alkynylgroup includes alkynyl groups in which a non-carbon or non-hydrogen atomis bonded to a carbon triple bonded to another carbon and those in whicha non-carbon or non-hydrogen atom is bonded to a carbon not involved ina triple bond to another carbon.

“Alkylene” by itself or as part of another substituent means a divalentradical derived from an alkane, as exemplified by —CH₂CH₂CH₂CH₂—.Typically, an alkylene group will have from 1, 2, 3, 4, 5, 6, 7 or 8carbon atoms that is derived by the removal of one hydrogen atom from asingle carbon atom of a parent alkyl.

“Cycloalkyl” or “carbocycle”, by themselves or in combination with otherterms, represent, unless otherwise stated, cyclic versions of “alkyl”,“alkenyl” and “alkynyl” in which all ring atoms are carbon. “Cycloalkyl”or “carbocycle” refers to a mono- or polycyclic group. When used inconnection with cycloalkyl substituents, the term “polycyclic” refersherein to fused and non-fused alkyl cyclic structures. “Cycloalkyl” or“carbocycle” may form a bridged ring or a spiro ring. The cycloalkylgroup may have one or more double or triple bond(s). The term“cycloalkenyl” refers to a cycloalkyl group that has at least one siteof alkenyl unsaturation between the ring vertices. The term“cycloalkynyl” refers to a cycloalkyl group that has at least one siteof alkynyl unsaturation between the ring vertices. When “cycloalkyl” isused in combination with “alkyl”, as in C₃₋₈cycloalkylC₃₋₈alkylene-, thecycloalkyl portion is meant to have the stated number of carbon atoms(e.g., from three to eight carbon atoms), while the alkylene portion hasfrom one to eight carbon atoms. Typical cycloalkyl substituents havefrom 3 to 8 ring atoms. Examples of cycloalkyl include cyclopentyl,cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like.

“Aryl” by itself or as part of another substituent refers to apolyunsaturated, aromatic, hydrocarbon group containing from 6 to 14carbon atoms, which can be a single ring or multiple rings (up to threerings) which are fused together or linked covalently. Thus the phraseincludes, but is not limited to, groups such as phenyl, biphenyl,anthracenyl, naphthyl by way of example. Non-limiting examples ofunsubstituted aryl groups include phenyl, 1-naphthyl, 2-naphthyl and4-biphenyl. “Substituted aryl group” includes, for example, —CH₂OH (onecarbon atom and one heteroatom replacing a carbon atom) and —CH₂SH. Theterm “heteroalkylene” by itself or as part of another substituent meansa divalent radical derived from heteroalkyl, as exemplified by—CH₂—CH₂—S—CH₂CH₂— and —CH₂—S—CH₂—CH₂—NH—CH₂—. For heteroalkylenegroups, heteroatoms can also occupy either or both of the chain termini(e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, andthe like). Still further, for alkylene and heteroalkylene linkinggroups, no orientation of the linking group is implied.

The terms “heterocycle”, “heterocyclyl” or “heterocyclic” refer to asaturated or unsaturated non-aromatic cyclic group containing at leastone heteroatom. As used herein, the term “heteroatom” is meant toinclude oxygen (O), nitrogen (N), sulfur (S) and silicon (Si). Eachheterocycle can be attached at any available ring carbon or heteroatom.Each heterocycle may have one or more rings. When multiple rings arepresent, they can be fused together or linked covalently. Eachheterocycle typically contains 1, 2, 3, 4 or 5, independently selectedheteroatoms. Preferably, these groups contain 1, 2, 3, 4, 5, 6, 7, 8, 9or 10 carbon atoms, 0, 1, 2, 3, 4 or 5 nitrogen atoms, 0, 1 or 2 sulfuratoms and 0, 1 or 2 oxygen atoms. More preferably, these groups contain1, 2 or 3 nitrogen atoms, 0-1 sulfur atoms and 0-1 oxygen atoms.Non-limiting examples of heterocycle groups include morpholin-3-one,piperazine-2-one, piperazin-1-oxide, pyridine-2-one, piperidine,morpholine, piperazine, isoxazoline, pyrazoline, imidazoline,pyrazol-5-one, pyrrolidine-2,5-dione, imidazolidine-2,4-dione,pyrrolidine, tetrahydroquinolinyl, decahydroquinolinyl,tetrahydrobenzooxazepinyl dihydrodibenzooxepin and the like.

“Heteroaryl” refers to a cyclic or polycyclic aromatic radical thatcontain from one to five heteroatoms selected from N, O, and S, whereinthe nitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A heteroaryl group can be attachedto the remainder of the molecule through a heteroatom or through acarbon atom and can contain 5 to 10 carbon atoms. Non-limiting examplesof heteroaryl groups include 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl,1-pyrazolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl,2-oxazolyl, 4-oxazolyl, 5-oxazolyl, 3-isoxazolyl, 4-isoxazolyl,5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl,2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl and4-pyrimidyl. If not specifically stated, substituents for each of theabove noted aryl and heteroaryl ring systems are selected from the groupof acceptable substituents described herein. “Substituted heteroaryl”refers to a unsubstituted heteroaryl group as defined above in which oneor more of the ring members is bonded to a non-hydrogen atom such asdescribed above with respect to substituted alkyl groups and substitutedaryl groups. Representative substituents include straight and branchedchain alkyl groups-CH₃, —C₂H₅, —CH₂OH, —OH, —OCH₃, —OC₂H₅, —OCF₃,—OC(═O)CH₃, —OC(═O)NH₂, —OC(═O)N(CH₃)₂, —CN, —NO₂, —C(═O)CH₃, —CO₂H,—CO₂CH₃, —CONH₂, —NH₂, —N(CH₃)₂, —NHSO₂CH₃, —NHCOCH₃, —NHC(═O)OCH₃,—NHSO₂CH₃, —SO₂CH₃, —SO₂NH₂ and halo.

“Bicyclic heteroaryl” refers to bicyclic aromatic radical that containfrom one to five heteroatoms selected from N, O, and S, wherein thenitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A bicyclic heteroaryl group can beattached to the remainder of the molecule through a heteroatom orthrough a carbon atom and can contain 5 to 10 carbon atoms. Non-limitingexamples of bicyclic heteroaryl groups include 5-benzothiazolyl,purinyl, 2-benzimidazolyl, benzopyrazolyl, 5-indolyl, azaindole,1-isoquinolyl, 5-isoquinolyl, 2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyland 6-quinolyl. If not specifically stated, substituents for each of theabove noted aryl and heteroaryl ring systems are selected from the groupof acceptable substituents described herein.

“Arylheteroaryl” or “aryleneheteroaryl” refers to a radical wherein aheteroaryl moiety is attached to an aryl moiety in a non-fused fashion.The aryl group is attached to the remainder of the molecule through acarbon atom and can contain 5 to 10 carbon atoms. Non-limiting examplesof aryl and heteroaryl groups are described above. The term“phenylheteraryl” or “phenyleneheteroaryl” refers to a heteroaryl moietyattached to a phenyl moiety which is attached to the remainder of themolecule.

“Arylheterocyclyl” or “aryleneheterocyclyl” refers to a radical whereina heterocyclyl moiety is attached to an aryl moiety in a non-fusedfashion. The aryl group is attached to the remainder of the moleculethrough a carbon atom and can contain 5 to 10 carbon atoms. Non-limitingexamples of aryl and heterocyclyl groups are described above. The term“phenylheterocyclyl” or “phenyleneheterocyclyl” refers to a heteroarylmoiety attached to a phenyl moiety which is attached to the remainder ofthe molecule.

In each of the above embodiments designating a number of atoms e.g.“C₁₋₈” is meant to include all possible embodiments that have one feweratom. Non-limiting examples include C₁₋₇, C₂₋₈, C₂₋₇, C₃₋₈, C₃₋₇ and thelike.

Each of the terms herein (e.g., “alkyl,” “heteroalkyl,” “aryl” and“heteroaryl”) is meant to include both “unsubstituted” and optionally“substituted” forms of the indicated radical, unless otherwiseindicated. Typically each radical is substituted with 0, 1, 2 3 4 or 5substituents, unless otherwise indicated. Examples of substituents foreach type of radical are provided below.

“Substituted” refers to a group as defined herein in which one or morebonds to a carbon(s) or hydrogen(s) are replaced by a bond tonon-hydrogen and non-carbon atom “substituents” such as, but not limitedto, a halogen atom such as F, Cl, Br, and I; an oxygen atom in groupssuch as hydroxyl groups, alkoxy groups, aryloxy, and acyloxy groups; asulfur atom in groups such as thiol groups, alkyl and aryl sulfidegroups, sulfone groups, sulfonyl groups, and sulfoxide groups; anitrogen atom in groups such as amino, alkylamines, dialkylamines,arylamines, alkylarylamines, diarylamines, alkoxyamino, hydroxyamino,acylamino, sulfonylamino, N-oxides, imides, and enamines; and otherheteroatoms in various other groups. “Substituents” also include groupsin which one or more bonds to a carbon(s) or hydrogen(s) atom isreplaced by a higher-order bond (e.g., a double- or triple-bond) to aheteroatom such as oxygen in oxo, acyl, amido, alkoxycarbonyl,aminocarbonyl, carboxyl, and ester groups; nitrogen in groups such asimines, oximes, hydrazones, and nitriles. “Substituents” further includegroups in which one or more bonds to a carbon(s) or hydrogen(s) atoms isreplaced by a bond to a cycloalkyl, heterocyclyl, aryl, and heteroarylgroups. Representative “substituents” include, among others, groups inwhich one or more bonds to a carbon or hydrogen atom is/are replaced byone or more bonds to fluoro, chloro, or bromo group. Anotherrepresentative “substituent” is the trifluoromethyl group and othergroups that contain the trifluoromethyl group. Other representative“substituents” include those in which one or more bonds to a carbon orhydrogen atom is replaced by a bond to an oxygen atom such that thesubstituted alkyl group contains a hydroxyl, alkoxy, or aryloxy group.Other representative “substituents” include alkyl groups that have anamine, or a substituted or unsubstituted alkylamine, dialkylamine,arylamine, (alkyl)(aryl)amine, diarylamine, heterocyclylamine,diheterocyclylamine, (alkyl)(heterocyclyl)amine, or(aryl)(heterocyclyl)amine group. Still other representative“substituents” include those in which one or more bonds to a carbon(s)or hydrogen(s) atoms is replaced by a bond to an alkyl, cycloalkyl,aryl, heteroaryl, or heterocyclyl group.

The herein-defined groups may include prefixes and/or suffixes that arecommonly used in the art to create additional well-recognizedsubstituent groups. As examples, “alkylamino” refers to a group of theformula —NR^(a)R^(b). Unless stated otherwise, for the following groupscontaining R^(a), R^(b), R^(c), R^(d) and R^(e): R^(a), and R^(b) areeach independently selected from H, alkyl, alkoxy, thioalkoxy,cycloalkyl, aryl, heteroaryl, or heterocyclyl or are optionally joinedtogether with the atom(s) to which they are attached to form a cyclicgroup. When R^(a) and R^(b) are attached to the same nitrogen atom, theycan be combined with the nitrogen atom to form a 5-, 6- or 7-memberedring. For example, —NR^(a)R^(b) is meant to include 1-pyrrolidinyl and4-morpholinyl.

R^(c), R^(d), R^(e) and R^(f) are each independently selected fromalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl,heterocyclyl or alkylenearyl as defined herein.

Typically, a particular radical will have 0, 1, 2 or 3 substituents,with those groups having two or fewer substituents being preferred inthe present invention. More preferably, a radical will be unsubstitutedor monosubstituted. Most preferably, a radical will be unsubstituted.

“Substituents” for the alkyl and heteroalkyl radicals (as well as thosegroups referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl,alkynyl, cycloalkyl, heterocyclyl) can be a variety of groups selectedfrom: —OR^(a), ═O, ═NR^(a), ═N—OR^(a), —NR^(a)R^(b), —SR^(a), halogen,—SiR^(a)R^(b)R^(c), —OC(O)R^(a), —C(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(a)—C(O)NR^(b)R^(c),—NR^(a)—SO₂NR^(b)R^(c), —NR^(b)CO₂R^(a), —NH—C(NH₂)═NH,—NR^(a)C(NH₂)═NH, —NH—C(NH₂)═NR^(a), —S(O)R^(a), —SO₂R^(a),—SO₂NR^(a)R^(b), —NR^(b)SO₂R, —CN and —NO₂, in a number ranging fromzero to three, with those groups having zero, one or two substituentsbeing particularly preferred.

In some embodiments, “substituents” for the alkyl and heteroalkylradicals are selected from: —OR^(a), ═O, —NR^(a)R^(b), —SR^(a), halogen,—SiR^(a)R^(b)R^(c), —OC(O)R^(a), —C(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(b)CO₂R^(a),—NR^(a)—SO₂NR^(b)R^(c), —S(O)R^(a), —SO₂R^(a), —SO₂NR^(a)R^(b),—NR^(c)SO₂R, —CN and —NO₂, where R^(a) and R^(b) are as defined above.In some embodiments, substituents are selected from: —OR^(a), ═O,—NR^(a)R^(b), halogen, —OC(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(b)CO₂R^(a),—NR^(a)—SO₂NR^(b)R^(c), —SO₂R^(a), —SO₂NR^(a)R^(b), —NR″SO₂R, —CN and—NO₂.

Examples of substituted alkyl are: —(CH₂)₃NH₂, —(CH₂)₃NH(CH₃),—(CH₂)₃NH(CH₃)₂, —CH₂C(═CH₂)CH₂NH₂, —CH₂C(═O)CH₂NH₂, —CH₂S(═O)₂CH₃,—CH₂OCH₂NH₂, —CO₂H. Examples of substituents of substituted alkyl are:CH₂OH, —OH, —OCH₃, —OC₂H₅, —OCF₃, —OC(═O)CH₃, —OC(═O)NH₂,—OC(═O)N(CH₃)₂, —CN, —NO₂, —C(═O)CH₃, —CO₂H, —CO₂CH₃, —CONH₂, —NH₂,—N(CH₃)₂, —NHSO₂CH₃, —NHCOCH₃, —NHC(═O)OCH₃, —NHSO₂CH₃, —SO₂CH₃,—SO₂NH₂, and halo.

Similarly, “substituents” for the aryl and heteroaryl groups are variedand are selected from: -halogen, —OR^(a), —OC(O)R^(a), —NR^(a)R^(b),—CN, —NO₂, —CO₂R^(a), —CONR^(a)R^(b), —C(O)R^(a), —OC(O)NR^(a)R^(b),—NR^(b)C(O)R^(a), —NR^(b)C(O)₂R^(a), —NR^(a)—C(O)NR^(b)R^(c),—NH—C(NH₂)═NH, —NR^(a)C(NH₂)═NH, —NH—C(NH₂)═NR^(a), —S(O)R^(a),—S(O)₂R^(a), —S(O) 2NR^(a)R^(b), —N₃, —CH(Ph)₂, perfluoroC₁₋₈alkoxy, andperfluoroC₁₋₈alkyl, in a number ranging from zero to the total number ofopen valences on the aromatic ring system; and where R^(a), R^(b) andR^(c) are independently selected from hydrogen, C₁₋₆alkyl andheteroalkyl, unsubstituted aryl and heteroaryl, (unsubstitutedaryl)-C₁₋₈alkyl, and (unsubstituted aryl)oxy-C₁₋₈alkyl.

Two of the “substituents” on adjacent atoms of the aryl or heteroarylring may optionally be replaced with a substituent of the formula-T-C(O)—(CH₂)q-U—, wherein T and U are independently —NH—, —O—, —CH_(2—)or a single bond, and q is 0, 1 or 2. Alternatively, two of thesubstituents on adjacent atoms of the aryl or heteroaryl ring mayoptionally be replaced with a substituent of the formula-A-(CH₂)_(r—)B—, wherein A and B are independently —CH_(2—), —O—, —NH—,—S—, —S(O)—, —S(O)_(2—), —S(O)₂NR^(a)— or a single bond, and r is 1, 2or 3. One of the single bonds of the new ring so formed may optionallybe replaced with a double bond. Alternatively, two of the substituentson adjacent atoms of the aryl or heteroaryl ring may optionally bereplaced with a substituent of the formula —(CH₂)_(s—)X—(CH₂)_(t—)—,where s and t are independently integers of 0 to 3, and X is —O—,—NR^(a)—, —S—, —S(O)—, —S(O)_(2—), or —S(O)₂NR^(a)—. The substituentR^(a) in —NR^(a)— and —S(O)₂NR^(a)— is selected from hydrogen orunsubstituted C₁₋₆alkyl. Otherwise, R′ is as defined above.

Unless indicated otherwise, the nomenclature of substituents that arenot explicitly defined herein are arrived at by naming the terminalportion of the functionality followed by the adjacent functionalitytoward the point of attachment. For example, the substituent“arylalkyloxycarbonyl” refers to the group (aryl)-(alkyl)-O—C(O)—.

The term “acyl” refers to the group —C(═O)R^(c) where R^(c) is alkyl,alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl orheterocyclyl. Acyl includes the “acetyl” group —C(═O)CH₃.

“Acylamino-” refers to the group —NR^(a)C(═O)R^(c) where R^(c) is alkyl,alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl orheterocyclyl.

“Acyloxy” refers to —OC(═O)—R^(c) where R^(c) is alkyl, alkenyl,alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl or heterocyclyl.

“Alkoxy” refers to —OR^(d) wherein R^(d) is alkyl as defined herein.Representative examples of alkoxy groups include methoxy, ethoxy,t-butoxy, trifluoromethoxy, and the like.

“Alkoxyamino” refers to the group —NHOR^(d) where R^(d) is alkyl.

“Alkoxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) is alkyl.Representative alkoxycarbonyl groups include, for example, those shownbelow.

These alkoxycarbonyl groups can be further substituted as will beapparent to those having skill in the organic and medicinal chemistryarts in conjunction with the disclosure herein.

“Alkoxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d) isalkyl.

“Alkoxysulfonylamino” refers to the group —NR^(a)S(═O)₂—OR^(d) whereR^(d) is alkyl.

“Alkylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) is alkyl.

“Alkylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) is alkyl.

“Alkylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) is alkyl.Representative alkylcarbonylamino groups include, for example,—NHC(═O)CH₃, —NHC(═O)CH₂CH₃, —NHC(═O)CH₂NH(CH₃), —NHC(═O)CH₂N(CH₃)₂, or—NHC(═O)(CH₂)₃OH.

“Alkylsulfanyl”, “alkylthio”, or “thioalkoxy” refers to the groupS—R^(d). where R^(d) is alkyl.

“Alkylsulfonyl” refers to —S(═O)₂R^(e) where R^(e) is alkyl.Alkylsulfonyl groups employed in compounds of the present invention aretypically C₁₋₆alkylsulfonyl groups.

“Alkylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e) isalkyl.

“Alkynyloxy” refers to the group —O-alkynyl, wherein alkynyl is asdefined herein. Alkynyloxy includes, by way of example, ethynyloxy,propynyloxy, and the like.

“Amidino” refers to the group —C(═NR^(a))NR^(b)R^(c), wherein R^(b) andR^(c) independently are selected from the group consisting of hydrogen,alkyl, substituted alkyl, alkenyl, alkynyl, aryl, cycloalkyl,cycloalkenyl, heteroaryl, heterocyclic, and where R^(b) and R^(c) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group. R^(a) is selected fromthe group consisting of hydrogen, alkyl, alkenyl, alkynyl, cycloalkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, substitutedheterocyclic, nitro, nitroso, hydroxy, alkoxy, cyano, —N═N—N-alkyl,—N(alkyl)SO₂-alkyl, —N═N═N-alkyl, acyl and —SO₂-alkyl.

“Amino” refers to a monovalent radical —NR^(a)R^(b) or divalent radical—NR^(a)—. The term “alkylamino” refers to the group —NR^(a)R^(b) whereR^(a) is alkyl and R^(b) is H or alkyl. The term “arylamino” refers tothe group —NR^(a)R^(b) where at least one R^(a) or R^(b) is aryl. Theterm “(alkyl)(aryl)amino” refers to the group —NR^(a)R^(b) where R^(a)is alkyl and R^(b) is aryl. Additionally, for dialkylamino groups, thealkyl portions can be the same or different and can also be combined toform a 3-7 membered ring with the nitrogen atom to which each isattached. Accordingly, a group represented as —NR^(a)R^(b) is meant toinclude piperidinyl, pyrrolidinyl, morpholinyl, azetidinyl and the like.

“Aminocarbonyl” or “aminoacyl” refers to the amide —C(═O—NR^(a)R^(b).The term “alkylaminocarbonyl” refers herein to the group—C(═O—NR^(a)R^(b) where R^(a) is alkyl and R^(b) is H or alkyl. The term“arylaminocarbonyl” refers herein to the group —C(═O—NR^(a)R^(b) whereR^(a) or R^(b) is aryl. Representative aminocarbonyl groups include, forexample, those shown below. These aminocarbonyl group can be furthersubstituted as will be apparent to those having skill in the organic andmedicinal chemistry arts in conjunction with the disclosure herein.

“Aminocarbonylamino” refers to the group —NR^(a)C(O)NR^(a)R^(b), whereinR^(a) is hydrogen or alkyl and R^(a) and R^(b) independently areselected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, and whereR^(a) and R^(b) are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminosulfonyl” refers to —S(O)₂NR^(a)R^(b) where R is independently areselected from the group consisting of hydrogen, alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl,substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and where R^(a) and R^(b) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group and alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl,cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Aminosulfonyloxy” refers to the group —O—SO₂NR^(a)R^(b), wherein R^(a)and R^(b) independently are selected from the group consisting ofhydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, cycloalkenyl,heteroaryl and heterocyclic; R^(a) and R^(b) are optionally joinedtogether with the nitrogen bound thereto to form a heterocyclic orsubstituted heterocyclic group.

“Aminosulfonylamino” refers to the group —NR^(a)—SO₂NR^(b)R^(c), whereinR^(a) is hydrogen or alkyl and R^(b) and R^(c) independently areselected from the group consisting of hydrogen, alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl,substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, heteroaryl, substituted heteroaryl,heterocyclic, and substituted heterocyclic and where R^(b) and R^(c) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group, and wherein alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Aminothiocarbonyl” refers to the group —C(S)NR^(a)R^(b), wherein R^(a)and R^(b) independently are selected from the group consisting ofhydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl,alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl,substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl,heteroaryl, substituted heteroaryl, heterocyclic, and substitutedheterocyclic and where R^(a) and R^(b) are optionally joined togetherwith the nitrogen bound thereto to form a heterocyclic or substitutedheterocyclic group, and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, andsubstituted heterocyclic are as defined herein.

“Aminothiocarbonylamino” refers to the group—NR^(a)C(S)NR^(a)R^(b),wherein R^(a) is hydrogen or alkyl and R^(b) and R^(c) are optionallyjoined together with the nitrogen bound thereto to form a heterocyclicor substituted heterocyclic group.

“Arylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) is aryl.

“Arylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) is aryl.

“Arylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) is aryl.

“Aryloxy” refers to —OR^(d) where R^(d) is aryl. Representative examplesof aryloxy groups include phenoxy, naphthoxy, and the like.

“Aryloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) is aryl.

“Aryloxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d) isaryl.

“Arylsulfanyl”, “arylthio”, or “thioaryloxy” refers to the groupS—R^(d). where R^(d) is aryl.

“Arylsulfonyl” refers to —S(═O)₂R^(e) where R^(e) is aryl.

“Arylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e) is aryl.

“Arylthio” refers to the group —S-aryl, wherein aryl is as definedherein. In other embodiments, sulfur may be oxidized to —S(O)— or —SO₂—moieties. The sulfoxide may exist as one or more stereoisomers.

“Azido” refers to —N₃.

“Bond” when used a element in a Markush group means that thecorresponding group does not exist, and the groups of both sides aredirectly linked.

“Carbonyl” refers to the divalent group —C(═O)—.

“Carboxy” or “carboxyl” refers to the group —CO₂H.

“Carboxyl ester” or “carboxy ester” refers to the groups —C(═O)OR^(c).

“(Carboxyl ester)amino” refers to the groups —NR^(a)—C(O)OR^(c), whereR^(a) is alkyl or hydrogen.

“(Carboxyl ester)oxy” or “Carbonate ester” refers to the groups—O—C(═O)OR^(c).

“Cyano” refers to —CN.

“Cycloalkoxy” refers to —OR^(d) where R^(d) is cycloalkyl.

“Cycloalkoxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) iscycloalkyl.

“Cycloalkoxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d) iscycloalkyl.

“Cycloalkylalkylene” refers to a radical —R^(x)R^(y) wherein R^(x) is analkylene group and R^(y) is a cycloalkyl group as defined herein, e.g.,cyclopropylmethyl, cyclohexenylpropyl, 3-cyclohexyl-2-methylpropyl, andthe like.

“Cycloalkylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) iscycloalkyl.

“Cycloalkylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) iscycloalkyl.

“Cycloalkylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) iscycloalkyl.

“Cycloalkylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(c) wherein R^(c) iscycloalkyl.

“Cycloalkylthio” refers to —S-cycloalkyl. In other embodiments, sulfurmay be oxidized to —S(O)— or —SO₂— moieties. The sulfoxide may exist asone or more stereoisomers.

“Cycloalkenylox” refers to —O-cycloalkenyl.

“Cycloalkenylthio” refers to —S-cycloalkenyl. In other embodiments,sulfur may be oxidized to sulfinyl or sulfonyl moieties. The sulfoxidemay exist as one or more stereoisomers.

“Ester” refers to —C(═O)OR^(d) wherein R^(d) is alkyl, cycloalkyl, aryl,heteroaryl, or heterocyclyl.

“Guanidino” refers to the group —NHC(═NH)NH₂.

“Halo” or “halogen” by themselves or as part of another substituent,mean, unless otherwise stated, a fluorine, chlorine, bromine, or iodineatom. Additionally, terms such as “haloalkyl”, are meant to includealkyl in which one or more hydrogen is substituted with halogen atomswhich can be the same or different, in a number ranging from one up tothe maximum number of halogens permitted e.g. for alkyl, (2m′+1), wherem′ is the total number of carbon atoms in the alkyl group. For example,the term “haloC₁₋₈ alkyl” is meant to include trifluoromethyl,2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like. Theterm “perhaloalkyl” means, unless otherwise stated, alkyl substitutedwith (2m′+1) halogen atoms, where m′ is the total number of carbon atomsin the alkyl group. For example, the term “perhaloC₁₋₈alkyl”, is meantto include trifluoromethyl, pentachloroethyl,1,1,1-trifluoro-2-bromo-2-chloroethyl, and the like. Additionally, term“haloalkoxy” refers to an alkoxy radical substituted with one or morehalogen atoms.

“Heteroalkyl” means an alkyl radical as defined herein with one, two orthree substituents independently selected from cyano, —OR^(w),—NR^(x)R^(y), and —S(O)_(n)R^(z) (where n is an integer from 0 to 2),with the understanding that the point of attachment of the heteroalkylradical is through a carbon atom of the heteroalkyl radical. R^(w) ishydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl,alkoxycarbonyl, aryloxycarbonyl, carboxamido, or mono- ordi-alkylcarbamoyl. R^(x) is hydrogen, alkyl, cycloalkyl,cycloalkyl-alkyl, aryl or araalkyl. R^(y) is hydrogen, alkyl,cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl,aryloxycarbonyl, carboxamido, mono- or di-alkylcarbamoyl oralkylsulfonyl. R^(z) is hydrogen (provided that n is 0), alkyl,cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl, amino, mono-alkylamino,di-alkylamino, or hydroxyalkyl. Representative examples include, forexample, 2-hydroxyethyl, 2,3-dihydroxypropyl, 2-methoxyethyl,benzyloxymethyl, 2-cyanoethyl, and 2-methylsulfonyl-ethyl. For each ofthe above, R^(w), R^(x), R^(y), and R^(z) can be further substituted byamino, fluorine, alkylamino, di-alkylamino, OH or alkoxy. Additionally,the prefix indicating the number of carbon atoms (e.g., C₁-C₁₀) refersto the total number of carbon atoms in the portion of the heteroalkylgroup exclusive of the cyano, —ORS', —NR^(x)R^(y), or —S(O)_(n)R^(z)portions.

“Heteroarylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) isheteroaryl.

“Heteroarylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) isheteroaryl.

“Heteroarylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) isheteroaryl.

“Heteroaryloxy” refers to —OR^(d) where R^(d) is heteroaryl.

“Heteroaryloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) isheteroaryl.

“Heteroaryloxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d)is heteroaryl.

“Heteroarylsulfonyllamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e)is heteroaryl.

“Heteroarylthio” refers to the group—S-heteroaryl. In other embodiments,sulfur may be oxidized to —S(O)— or —SO₂— moieties. The sulfoxide mayexist as one or more stereoisomers.

“Heterocyclylalkyl” or “Cycloheteroalkyl-alkyl” means a radical—R^(x)R^(y) where R^(x) is an alkylene group and R^(y) is a heterocyclylgroup as defined herein, e.g., tetrahydropyran-2-ylmethyl,4-(4-substituted-phenyl)piperazin-1-ylmethyl, 3-piperidinylethyl, andthe like.

“Heterocycloxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d)is heterocyclyl.

“Heterocyclylcarbonyl” refers to the —C(═O)R^(c) where R^(c) isheterocyclyl.

“Heterocyclylcarbonylamino” refers to —NR^(a)C(—O)R^(c) wherein R^(c) isheterocyclyl.

“Heterocyclylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) sheterocyclyl.

“Heterocyclyloxy” refers to —OR^(d) where R^(d) is heterocyclyl.

“Heterocyclyloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) isheterocyclyl.

“Heterocyclylsulfonyl” refers to —S(═O)₂R^(e) where R^(e) isheterocyclyl.

“Heterocyclylsulfonyllamino” refers to —NR^(b)S(═O)₂—R^(c) wherein R^(c)is heterocyclyl.

“Heterocyclylthio” refers to the group —S-heterocycyl. In otherembodiments, sulfur may be oxidized to —S(O)— or —SO₂— moieties. Thesulfoxide may exist as one or more stereoisomers.

“Hydroxy” or “hydroxyl” refers to the group —OH.

“Hydroxyamino” refers to the group —NHOH.

“Nitro” refers to —NO₂.

“Nitroso” refers to the group —NO.

The terms “optional” or “optionally” as used throughout thespecification means that the subsequently described event orcircumstance may but need not occur, and that the description includesinstances where the event or circumstance occurs and instances in whichit does not. For example, “heterocyclo group optionally mono- ordi-substituted with an alkyl group means that the alkyl may but need notbe present, and the description includes situations where theheterocyclo group is mono- or disubstituted with an alkyl group andsituations where the heterocyclo group is not substituted with the alkylgroup.

“Optionally substituted” means a ring which is optionally substitutedindependently with substituents. A site of a group that is unsubstitutedmay be substituted with hydrogen.

“Oxo” refers to the divalent group ═O.

“Sulfanyl” refers to the group —SR^(f) where R^(f) is as defined herein.

“Sulfinyl” refers to the group —S(═O)—R^(e) where R^(e) is as definedherein.

“Sulfonic acid” refers to the group —S(O)₂—OH.

“Sulfonyl” refers to the group —S(O)₂—R^(e) where R^(e) is as definedherein.

“Sulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) where R^(a) is selectedfrom the group consisting of hydrogen, alkyl, alkenyl, alkynyl, aryl,cycloalkyl, cycloalkenyl, heteroaryl and heterocyclyl and R^(e) is asdefined herein.

“Sulfonyloxy” refers to the group —OSO₂—R^(c).

Compounds that have the same molecular formula but differ in the natureor sequence of bonding of their atoms or the arrangement of their atomsin space are termed “isomers”. Isomers that differ in the arrangement oftheir atoms in space are termed “stereoisomers”. “Stereoisomer” and“stereoisomers” refer to compounds that exist in differentstereoisomeric forms if they possess one or more asymmetric centers or adouble bond with asymmetric substitution and, therefore, can be producedas individual stereoisomers or as mixtures. Stereoisomers includeenantiomers and diastereomers. Stereoisomers that are not mirror imagesof one another are termed “diastereomers” and those that arenon-superimposable mirror images of each other are termed “enantiomers”.When a compound has an asymmetric center, for example, it is bonded tofour different groups, a pair of enantiomers is possible. An enantiomercan be characterized by the absolute configuration of its asymmetriccenter and is described by the R- and S-sequencing rules of Calm andPrelog, or by the manner in which the molecule rotates the plane ofpolarized light and designated as dextrorotatory or levorotatory (i.e.,as (+) or (−)-isomers respectively). A chiral compound can exist aseither individual enantiomer or as a mixture thereof. A mixturecontaining equal proportions of the enantiomers is called a “racemicmixture”. Unless otherwise indicated, the description is intended toinclude individual stereoisomers as well as mixtures. The methods forthe determination of stereochemistry and the separation of stereoisomersare well-known in the art (see discussion in Chapter 4 of ADVANCEDORGANIC CHEMISTRY, 4th edition J. March, John Wiley and Sons, New York,1992) differ in the chirality of one or more stereocenters.

“Thioacyl” refers to the groups R^(a)—C(S)—.

“Thiol” refers to the group—SH.

“Tautomer” refers to alternate forms of a molecule that differ in theposition of a proton, such as enol-keto and imine-enamine tautomers, orthe tautomeric forms of heteroaryl groups containing a —N═C(H)—NH— ringatom arrangement, such as pyrazoles, imidazoles, benzimidazoles,triazoles, and tetrazoles. A person of ordinary skill in the art wouldrecognize that other tautomeric ring atom arrangements are possible.

It is understood that in all substituted groups defined above, polymersarrived at by defining substituents with further substituents tothemselves (e.g., substituted aryl having a substituted aryl group as asubstituent which is itself substituted with a substituted aryl group,which is further substituted by a substituted aryl group, etc.) are notintended for inclusion herein. In such cases, the maximum number of suchsubstitutions is three. For example, serial substitutions of substitutedaryl groups are limited to -substituted aryl-(substitutedaryl)-substituted aryl.

“Protecting group” refers to a group of atoms that, when attached to areactive functional group in a molecule, mask, reduce or prevent thereactivity of the functional group. Typically, a protecting group may beselectively removed as desired during the course of a synthesis.Examples of protecting groups can be found in Greene and Wuts,Protective Groups in Organic Chemistry, 3^(rd) Ed., 1999, John Wiley &Sons, NY and Harrison et al., Compendium of Synthetic Organic Methods,Vols. 1-8, 1971-1996, John Wiley & Sons, NY. Representative aminoprotecting groups include, but are not limited to, formyl, acetyl,trifluoroacetyl, benzyl, benzyloxycarbonyl (“CBZ”), tert-butoxycarbonyl(“Boc”), trimethylsilyl (“TMS”), 2-trimethylsilyl-ethanesulfonyl(“TES”), trityl and substituted trityl groups, allyloxycarbonyl,9-fluorenylmethyloxycarbonyl (“FMOC”), nitro-veratryloxycarbonyl(“NVOC”) and the like. Representative hydroxy protecting groups include,but are not limited to, those where the hydroxy group is either acylatedor alkylated such as benzyl and trityl ethers, as well as alkyl ethers,tetrahydropyranyl ethers, trialkylsilyl ethers (e.g., TMS or TIPPSgroups) and allyl ethers.

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituents found on thecompounds described herein. When compounds of the present inventioncontain relatively acidic functionalities, base addition salts can beobtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of salts derived frompharmaceutically-acceptable inorganic bases include aluminum, ammonium,calcium, copper, ferric, ferrous, lithium, magnesium, manganic,manganous, potassium, sodium, zinc and the like. Salts derived frompharmaceutically-acceptable organic bases include salts of primary,secondary and tertiary amines, including substituted amines, cyclicamines, naturally-occurring amines and the like, such as arginine,betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperidine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, malonic, benzoic, succinic, suberic,fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric,tartaric, methanesulfonic, and the like. Also included are salts ofamino acids such as arginate and the like, and salts of organic acidslike glucuronic or galactunoric acids and the like (see, e.g., Berge, S.M. et al., “Pharmaceutical Salts,” Journal of Pharmaceutical Science,66:1-19, 1977). Certain specific compounds of the present inventioncontain both basic and acidic functionalities that allow the compoundsto be converted into either base or acid addition salts.

The neutral forms of the compounds may be regenerated by contacting thesalt with a base or acid and isolating the parent compound in theconventional manner. The parent form of the compound differs from thevarious salt forms in certain physical properties, such as solubility inpolar solvents, but otherwise the salts are equivalent to the parentform of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compoundswhich are in a prodrug ester form. “Prodrug”s of the compounds describedherein are those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent. Prodrugs arefrequently, but not necessarily, pharmacologically inactive untilconverted into the active drug. Prodrugs are typically obtained bymasking a functional group in the drug believed to be in part requiredfor activity with a progroup (defined below) to form a promoiety whichundergoes a transformation, such as cleavage, under the specifiedconditions of use to release the functional group, and hence the activedrug. The cleavage of the promoiety may proceed spontaneously, such asby way of a hydrolysis reaction, or it may be catalyzed or induced byanother agent, such as by an enzyme, by light, by acid or base, or by achange of or exposure to a physical or environmental parameter, such asa change of temperature. The agent may be endogenous to the conditionsof use, such as an enzyme present in the cells to which the prodrug isadministered or the acidic conditions of the stomach, or it may besupplied exogenously.

“Progroup” refers to a type of protecting group that, when used to maska functional group within an active drug to form a promoiety, convertsthe drug into a prodrug. Progroups are typically attached to thefunctional group of the drug via bonds that are cleavable underspecified conditions of use. Thus, a progroup is that portion of apromoiety that cleaves to release the functional group under thespecified conditions of use. As a specific example, an amide promoietyof the formula —NH—C(O)CH₃ comprises the progroup —C(O)CH₃.

A wide variety of progroups, as well as the resultant promoieties,suitable for masking functional groups in the active syk selectiveinhibitory compounds to yield prodrugs are well-known in the art. Forexample, a hydroxyl functional group may be masked as a sulfonate, ester(such as acetate or maleate) or carbonate promoiety, which may behydrolyzed in vivo to provide the hydroxyl group. An amino functionalgroup may be masked as an amide, carbamate, imine, urea, phosphenyl,phosphoryl or sulfenyl promoiety, which may be hydrolyzed in vivo toprovide the amino group. A carboxyl group may be masked as an ester(including methyl, ethyl, pivaloyloxymethyl, silyl esters andthioesters), amide or hydrazide promoiety, which may be hydrolyzed invivo to provide the carboxyl group. The invention includes those estersand acyl groups known in the art for modifying the solubility orhydrolysis characteristics for use as sustained-release or prodrugformulations. Other specific examples of suitable progroups and theirrespective promoieties will be apparent to those of skill in the art.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. “Solvate” refers toa complex formed by combination of solvent molecules with molecules orions of the solute. The solvent can be an organic compound, an inorganiccompound, or a mixture of both. Some examples of solvents include, butare not limited to, methanol, N,N-dimethylformamide, tetrahydrofuran,dimethylsulfoxide, and water. In general, the solvated forms areequivalent to unsolvated forms and are intended to be encompassed withinthe scope of the present invention. Certain compounds of the presentinvention may exist in multiple crystalline or amorphous forms. Ingeneral, all physical forms are equivalent for the uses contemplated bythe present invention and are intended to be within the scope of thepresent invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds; the racemates, diastereomers,geometric isomers, regioisomers and individual isomers (e.g., separateenantiomers) are all intended to be encompassed within the scope of thepresent invention. These isomers can be resolved or asymmetricallysynthesized using conventional methods to render the isomers “opticallypure”, i.e., substantially free of its other isomers. If, for instance,a particular enantiomer of a compound of the present invention isdesired, it may be prepared by asymmetric synthesis, or by derivationwith a chrial auxilliary, where the resulting diastereomeric mixture isseparated and the auxilliary group cleaved to provide the pure desiredenantiomers. Alternatively, where the molecule contains a basicfunctional group, such as amino, or an acidic functional group, such ascarboxyl, diastereomeric salts are formed with an appropriateoptically-active acid or base, followed by resolution of thediasteromers thus formed by fractional crystallization orchromatographic means well known in the art, and subsequent recovery ofthe pure enantiomers.

The compounds of the present invention may also contain unnaturalproportions of atomic isotopes at one or more of the atoms thatconstitute such compounds. For example, the compounds may beradiolabeled with radioactive isotopes, such as for example tritium(³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations ofthe compounds of the present invention, whether radioactive or not, areintended to be encompassed within the scope of the present invention.

The term “administering” refers to oral administration, administrationas a suppository, topical contact, intravenous, intraperitoneal,intramuscular, intralesional, intranasal or subcutaneous administration,or the implantation of a slow-release device e.g., a mini-osmotic pump,to a subject. Administration is by any route, including parenteral andtransmucosal (e.g., buccal, sublingual, palatal, gingival, nasal,vaginal, rectal, or transdermal). Parenteral administration includes,e.g., intravenous, intramuscular, intra-arteriole, intradermal,subcutaneous, intraperitoneal, intraventricular, and intracranial. Othermodes of delivery include, but are not limited to, the use of liposomalformulations, intravenous infusion, transdermal patches, etc.

An “agonist” or “activator” refers to an agent or molecule that binds toa receptor of the invention, stimulates, increases, opens, activates,facilitates, enhances activation or enzymatic activity, sensitizes or upregulates the activity of a receptor of the invention.

An “antagonist” or “inhibitor” refers to an agent or molecule thatinhibits or binds to, partially or totally blocks stimulation oractivity, decreases, closes, prevents, delays activation or enzymaticactivity, inactivates, desensitizes, or down regulates the activity of areceptor of the invention. As used herein, “antagonist” also includes areverse or inverse agonist.

As used herein, the term “condition or disorder responsive to modulationof syk” and related terms and phrases refer to a condition or disorderassociated with inappropriate, e.g., less than or greater than normal,activity of syk and at least partially responsive to or affected bymodulation of syk (e.g., syk antagonist or agonist results in someimprovement in patient well-being in at least some patients).Inappropriate functional activity of syk might arise as the result ofexpression of syk in cells which normally do not express the receptor,greater than normal production of syk, or slower than normal metabolicinactivation or elimination of syk or its active metabolites, increasedexpression of syk or degree of intracellular activation (leading to,e.g., inflammatory and immune-related disorders and conditions) ordecreased expression of syk. A condition or disorder associated with sykmay include a “syk-mediated condition or disorder”.

As used herein, the phrases “a condition or disorder mediated at leastin part by syk kinase activity”, and related phrases and terms refer toa condition or disorder characterized by inappropriate, e.g., greaterthan normal, syk activity. Inappropriate syk functional activity mightarise as the result of syk expression in cells which normally do notexpress syk or increased syk expression or degree of intracellularactivation (leading to, e.g., inflammatory and immune-related disordersand conditions). A condition or disorder mediated at least in part bysyk kinase activity may be completely or partially mediated byinappropriate syk functional activity. However, a condition or disordermediated at least in part by syk kinase activity is one in whichmodulation of syk results in some effect on the underlying condition ordisorder (e.g., an syk antagonist results in some improvement in patientwell-being in at least some patients).

The term “inflammation” as used herein refers to infiltration of whiteblood cells (e.g., leukocytes, monocytes, etc.) into the area beingtreated for restenosis.

The term “intervention” refers to an action that produces an effect orthat is intended to alter the course of a disease process. For example,“vascular intervention” refers to the use of an intravascular proceduresuch as angioplasty or a stent to open an obstructed blood vessel.

The term “intravascular device” refers to a device useful for a vascularrecanalization procedure to restore blood flow through an obstructedblood vessel. Examples of intravascular devices include, withoutlimitation, stents, balloon catheters, autologous venous/arterialgrafts, prosthetic venous/arterial grafts, vascular catheters, andvascular shunts.

As used herein, the term “JAK” refers to a Janus kinase (RefSeqAccession No. P-43408) or a variant thereof that is capable of mediatinggene expression in vitro or in vivo. JAK variants include proteinssubstantially homologous to native JAK, i.e., proteins having one ormore naturally or non-naturally occurring amino acid deletions,insertions or substitutions (e.g., JAK derivatives, homologs andfragments). The amino acid sequence of JAK variant preferably is atleast about 80% identical to a native JAK, more preferably at leastabout 90% identical, and most preferably at least about 95% identical.

The term “leukocyte” refers to any of the various blood cells that havea nucleus and cytoplasm, separate into a thin white layer when wholeblood is centrifuged, and help protect the body from infection anddisease. Examples of leukocytes include, without limitation,neutrophils, eosinophils, basophils, lymphocytes, and monocytes.

The term “mammal” includes, without limitation, humans, domestic animals(e.g., dogs or cats), farm animals (cows, horses, or pigs), monkeys,rabbits, mice, and laboratory animals.

The terms “modulate”, “modulation” and the like refer to the ability ofa compound to increase or decrease the function and/or expression ofsyk, where such function may include transcription regulatory activityand/or protein-binding. Modulation may occur in vitro or in vivo.Modulation, as described herein, includes the inhibition, antagonism,partial antagonism, activation, agonism or partial agonism of a functionor characteristic associated with syk, either directly or indirectly,and/or the upregulation or downregulation of the expression of syk,either directly or indirectly. In a preferred embodiment, the modulationis direct. Inhibitors or antagonists are compounds that, e.g., bind to,partially or totally block stimulation, decrease, prevent, inhibit,delay activation, inactivate, desensitize, or downregulate signaltransduction. Activators or agonists are compounds that, e.g., bind to,stimulate, increase, open, activate, facilitate, enhance activation,activate, sensitize or upregulate signal transduction. The ability of acompound to inhibit the function of syk can be demonstrated in abiochemical assay, e.g., binding assay, or a cell-based assay, e.g., atransient transfection assay.

“Modulators” of activity are used to refer to “ligands”, “antagonists”and “agonists” identified using in vitro and in vivo assays for activityand their homologs and mimetics. Modulators include naturally occurringand synthetic ligands, antagonists, agonists, molecules and the like.Assays to identify antagonists and agonists include, e.g., applyingputative modulator compounds to cells, in the presence or absence of areceptor of the invention and then determining the functional effects ona receptor of the invention activity. Samples or assays comprising areceptor of the invention that are treated with a potential activator,inhibitor, or modulator are compared to control samples without theinhibitor, activator, or modulator to examine the extent of effect.Control samples (untreated with modulators) are assigned a relativeactivity value of 100%. Inhibition is achieved when the activity valueof a receptor of the invention relative to the control is about 80%,optionally 50% or 25-1%. Activation is achieved when the activity valueof a receptor of the invention relative to the control is 110%,optionally 150%, optionally 200-500%, or 1000-3000% higher.

“Patient” refers to human and non-human animals, especially mammals.Examples of patients include, but are not limited to, humans, cows,dogs, cats, goats, sheep, pigs and rabbits.

Turning next to the compositions of the invention, the term“pharmaceutically acceptable carrier or excipient” means a carrier orexcipient that is useful in preparing a pharmaceutical composition thatis generally safe, non-toxic and neither biologically nor otherwiseundesirable, and includes a carrier or excipient that is acceptable forveterinary use as well as human pharmaceutical use. A “pharmaceuticallyacceptable carrier or excipient” as used in the specification and claimsincludes both one and more than one such carrier or excipient.

The terms “pharmaceutically effective amount”, “therapeuticallyeffective amount” or “therapeutically effective dose” refers to theamount of the subject compound that will elicit the biological ormedical response of a tissue, system, animal or human that is beingsought by the researcher, veterinarian, medical doctor or otherclinician. The term “therapeutically effective amount” includes thatamount of a compound that, when administered, is sufficient to preventdevelopment of, or alleviate to some extent, one or more of the symptomsof the condition or disorder being treated. The therapeuticallyeffective amount will vary depending on the compound, the disorder orcondition and its severity and the age, weight, etc., of the mammal tobe treated.

The term “platelet” refers to a minute, normucleated, disklike cellfound in the blood plasma of mammals that functions to promote bloodclotting.

The terms “prevent”, “preventing”, “prevention” and grammaticalvariations thereof as used herein, refers to a method of partially orcompletely delaying or precluding the onset or recurrence of a disorderor condition and/or one or more of its attendant symptoms or barring asubject from acquiring or reacquiring a disorder or condition orreducing a subject's risk of acquiring or reaquiring a disorder orcondition or one or more of its attendant symptoms.

The term “recanalization” refers to the process of restoring flow to orreuniting an interrupted channel of the body, such as a blood vessel.

The term “restenosis” refers to a re-narrowing or blockage of an arteryat the same site where treatment, such as an angioplasty or a stentprocedure, has been performed.

The phrase “selectively” or “specifically” when referring to binding toa receptor, refers to a binding reaction that is determinative of thepresence of the receptor, often in a heterogeneous population ofreceptors and other biologics. Thus, under designated conditions, thecompounds bind to a particular receptor at least two times thebackground and more typically more than 10 to 100 times background.Specific binding of a compound under such conditions requires a compoundthat is selected for its specificity for a particular receptor. Forexample, small organic molecules can be screened to obtain only thosecompounds that specifically or selectively bind to a selected receptorand not with other receptors or proteins. A variety of assay formats maybe used to select compounds that are selective for a particularreceptor. For example, High-throughput screening assays are routinelyused to select compounds that are selective for a particular a receptor.

As used herein, the term “Sickle cell anemia” refers to an inheriteddisorder of the red blood cells in which both hemoglobin alleles encodethe sickle hemoglobin (S) protein, i.e., the S/S genotype. The presenceof abnormal hemoglobin results in the production of unusually shapedcells, which do not survive the usual length of time in the bloodcirculation. Thus, anemia results. “Anemia” refers to a decrease in thenumber of red blood cells and/or hemoglobin in the blood.

The term “Sickle cell disease” refers to an inherited disorder of thered blood cells in which one hemoglobin allele encodes the sicklehemoglobin (S) protein, and the other allele encodes another unusualhemoglobin protein, such as hemoglobin (S), (C), (D), (E), and (βThal).Examples of sickle cell disease genotypes include, without limitation,the S/S, S/C, S/D, S/E, and S/βThal genotypes. The most common types ofsickle cell disease include sickle cell anemia, sickle-hemoglobin Cdisease, sickle beta-plus thalassemia, and sickle beta-zero thalassemia.

The “subject” is defined herein to include animals such as mammals,including, but not limited to, primates (e.g., humans), cows, sheep,goats, horses, dogs, cats, rabbits, rats, mice and the like. Inpreferred embodiments, the subject is a human.

As used herein, the term “syk” refers to a spleen tyrosine kinase(RefSeq Accession No. P-043405) or a variant thereof that is capable ofmediating a cellular response to T-cell receptors in vitro or in vivo.syk variants include proteins substantially homologous to native syk,i.e., proteins having one or more naturally or non-naturally occurringamino acid deletions, insertions or substitutions (e.g., sykderivatives, homologs and fragments). The amino acid sequence of sykvariant preferably is at least about 80% identical to a native syk, morepreferably at least about 90% identical, and most preferably at leastabout 95% identical.

The term “syk inhibitor” refers to any agent that inhibits the catalyticactivity of spleen tyrosine kinase.

The term “thrombosis” refers to the blockage or clotting of a bloodvessel caused by a clumping of cells, resulting in the obstruction ofblood flow. The term “thrombosis” refers to the clot that is formedwithin the blood vessel.

The terms “treat”, “treating”, “treatment” and grammatical variationsthereof as used herein, includes partially or completely delaying,alleviating, mitigating or reducing the intensity of one or moreattendant symptoms of a disorder or condition and/or alleviating,mitigating or impeding one or more causes of a disorder or condition.Treatments according to the invention may be applied preventively,prophylactically, pallatively or remedially.

The term “vessel” refers to any channel for carrying a fluid, such as anartery or vein. For example, a “blood vessel” refers to any of thevessels through which blood circulates in the body. The lumen of a bloodvessel refers to the inner open space or cavity of the blood vessel.

2. Embodiments of the Invention

a. Compounds

The present invention provides in one group of embodiments, a compoundhaving formula (I):

or a tautomer thereof or a pharmaceutically acceptable salt thereof,wherein:

Y¹ is selected from the group consisting of:

Z is O or S;

D¹ is selected from the group consisting of:

-   -   (a) phenyl substituted with a group, R⁵, wherein the phenyl is        further optionally substituted with from 1 to 2 substituents,        R^(7a), independently selected from the group consisting of        C₁₋₈alkyl, C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and heterocyclyl;    -   R⁵ is selected from the group consisting of:        -   (i) heteroaryl;        -   (ii) heterocyclyl;        -   (iii) C₁₋₈alkylheterocyclyl;        -   (iv) phenyleneheteroaryl        -   (v) phenyleneheterocyclyl        -   (vi) -L-phenyl;        -   (vii) -L-heterocyclyl; and        -   (viii) acyloxy;    -   L is selected from the group consisting of —CO—, —O—, —SO₂—,        —CONH— and —CONHCH₂—;    -   each R⁵ is optionally further substituted with from 1 to 2        substituents independently selected from the group consisting of        C₁₋₈alkyl, hydroxyC₁₋₈alkyl-, aminoC₁₋₈alkyl, C₁₋₈alkylamino,        C₁₋₈alkylcarbonyl, aminocarbonyl, cyano, hydroxy, oxo, halo,        haloC₁₋₈alkyl, aminosulfonyl, C₃₋₈cycloalkyl and aryl;    -   (b) naphthyl substituted a substituent, R^(7b), selected from        the group consisting of halogen, C₁₋₈alkylcarbonyl,        C₁₋₈alkylsulfonyl, aminosulfonyl, heterocyclylcarbonyl and        aminocarbonyl;    -   (c) C₃₋₈cycloalkyl, optionally substituted with from 1 to 2        substituents, R^(7c), independently selected from the group        consisting of C₁₋₈alkyl, C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and        heterocyclyl;    -   (d) heteroaryl; optionally substituted with from 1 to 2        substituents, R^(7c), independently selected from the group        consisting of: C₁₋₈ alkyl, amino, hydroxyl, oxo, halo, C₁₋₈        alkoxy, hydroxyC₁₋₈alkyl-, aminoC₁₋₈alkyl, C₁₋₈alkylcarbonyl,        haloC₁₋₈alkyl, C₃₋₈cycloalkyl, C₁₋₈-aminocycloalkyl,        aminoC₁₋₈alkylenecarbonyl, aminocarbonyl,        C₁₋₈alkyleneaminoC₁₋₈alkylenecarbonyl,        C₁₋₈alkoxyC₁₋₈alkylenecarbonyl, hydroxyC₁₋₈alkylenecarbonyl,        hydroxyC₁₋₈alkoxycarbonyl, C₁₋₈ alkoxycarbonylamino, aryl,        arylC₁₋₈alkoxycarbonylamino, C₁₋₈alkylsulfonyl,        aminoC₁₋₈alkylenesulfonyl, aminosulfonyl,        C₁₋₈alkyleneaminoC₁₋₈alkylenesulfonyl,        C₁₋₈alkoxyC₁₋₈alkylenesulfonyl, hydroxyC₁₋₈alkylenesulfonyl,        hydroxyC₁₋₈alkoxysulfonyl, aminosulfonyl, and        C₁₋₈alkylheterocyclyl; and    -   (e) heterocyclyl; with from 1 to 2 substituents, R^(7e),        independently selected from the group consisting of C₁₋₈alkyl,        C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and heterocyclyl;

each E¹ is independently selected from the group consisting of C₁₋₈alkyl, C₂₋₈ alkenyl, C₂₋₈alkynyl, C₁₋₈alkoxy, C₁₋₈alkylthio,aminocarbonyl, C₁₋₈alkoxycarbonylC₁₋₈alkylene,C₁₋₈alkoxycarbonylC₁₋₈C₁₋₈alkoxy, C₁₋₈alkoxycarbonylamino, oxo, halo,cyano, haloC₁₋₈alkyl, haloC₁₋₈alkoxy, aminosulfonyl, heteroarylsulfinyl;amino, hydroxyl, C₁₋₈arylalkylene, phenyl, aminoC₁₋₈alkyl,aminoC₃₋₈cycloalkyl, heterocyclyl, heteroaryl andheterocyclylC₁₋₈alkylene;

each R^(1a), R^(1b) and R^(1c) is independently selected from the groupconsisting of: H, C₁₋₈alkyl, hydroxyC₁₋₈alkyl, C₁₋₈haloalkyl, amino,C₁₋₈alkylamino, C₁₋₈ alkoxycarbonylaminoC₁₋₈ alkylene, C₃₋₈cycloalkyl,heteroaryl, C₁₋₈ alkylC₃₋₈cycloalkyl, C₁₋₈alkylthioC₁₋₈ alkyl,C₁₋₈alkylsulfonylC₁₋₈ alkylene, aminocarbonyl, C₁₋₈alkoxyC₁₋₈alkyl,haloC₁₋₈alkyl, aryl and heterocyclyl; wherein the aryl is optionallysubstituted by hydroxyl, C₁₋₈alkoxy, halo or haloC₁₋₈alkyl; or takentogether with R³ and the atoms to which they are attached to form a C₃₋₈cycloalkyl or heterocycloalkyl ring;

R² is selected from the group consisting of H, amino, C₁₋₈alkylamino,hydroxycarbonylamino C₁₋₈alkoxycarbonylamino,arylC₁₋₈alkoxycarbonylamino and hydroxyl;

R³ is selected from the group consisting of H, C₁₋₈alkyl,C₁₋₈alkylamino, amino aminoC₁₋₈alkyl, carboxy, C₁₋₈alkylaminoC₁₋₈alkyl,C₁₋₈alkoxyC₁₋₈alkyl, hydroxyC₁₋₈alkyl; carboxyC₁₋₈alkyl,C₃₋₈cycloalkylC₁₋₈alkyl, aryloxyC₁₋₈alkyl, arylC₁₋₈alkyl,heteroarylC₁₋₈alkyl, and hydroxyC₁₋₈alkoxy and hydroxyC₁₋₈alkoxy; or maybe combined with R^(1c) or R⁴ and the atoms to which they are attachedto form a C₃₋₈ cycloalkyl or heterocyclyl ring;

R⁴ is H or alkyl or may be combined with R³ and the atoms to which theyare attached to form a C₃₋₈ cycloalkyl or heterocyclyl ring;

the subscript n is 0, 1, 2, 3 or 4; and

the subscript m is an integer of 1, 2 or 3; and the wavy line indicatesthe point of attachment to the rest of the molecule.

In one group of embodiments, Y¹ is:

In one group of embodiments, Y¹ is:

In one group of embodiments, Z is O. In another group of embodiments, Zis S.

In one group of embodiments, D¹ is phenyl.

In one group of embodiments, R⁵ is heteroaryl. In another group ofembodiments, R⁵ is heterocyclyl. In another group of embodiments, R⁵ isC₁₋₈alkylheterocyclyl. In another group of embodiments, R⁵ isphenyleneheteroaryl. In another group of embodiments, R⁵ isphenyleneheterocyclyl. In another group of embodiments, R⁵ is -L-phenyl.In another group of embodiments, R⁵ is -L-heterocyclyl. In another groupof embodiments, R⁵ is acyloxy.

In another group of embodiments, D¹ is naphthyl.

The present invention provides in another embodiment, a compound whereinD¹- is selected from the group consisting of:

wherein X is halogen.

The present invention provides in another embodiment, a compound whereinD¹- is selected from the group consisting of:

In another group of embodiments, D¹ is C₃₋₈cycloalkyl. In another groupof embodiments, D¹ is heteroaryl. In another group of embodiments, D¹ isheterocyclyl.

The present invention provides in another embodiment, a compound havingformula (I):

or a pharmaceutically acceptable salt thereof,wherein:

Y¹ is selected from the group consisting of:

D¹ is selected from the group consisting of:

-   -   (a) phenyl substituted with a heteroaryl group R⁵, wherein the        phenyl is further optionally substituted with from 1 to 2        substituents, R^(7a), independently selected from the group        consisting of C₁₋₈alkyl, C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and        heterocyclyl; and the heteroaryl is optionally further        substituted with from 1 to 2 substituents independently selected        from the group consisting of C₁₋₈alkyl, hydroxyC₁₋₈alkyl-,        aminoC₁₋₈alkyl, C₁₋₈alkylcarbonyl, aminocarbonyl, hydroxy, oxo,        halo, haloC₁₋₈alkyl, aminosulfonyl and C₃₋₈cycloalkyl;    -   (b) naphthyl substituted a substituent, R^(7b), selected from        the group consisting of C₁₋₈alkylsulfonyl, aminosulfonyl,        heterocyclylcarbonyl and aminocarbonyl;    -   (c) C₃₋₈cycloalkyl, optionally substituted with from 1 to 2        substituents, R^(7c), independently selected from the group        consisting of C₁₋₈alkyl, C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and        heterocyclyl;    -   (d) bicyclic heteroaryl; optionally substituted with from 1 to 2        substituents, R^(7d), independently selected from the group        consisting of: C₁₋₈ alkyl, C₁₋₈alkylcarbonyl,        aminoC₁₋₈alkylenecarbonyl, aminocarbonyl,        C₁₋₈alkyleneaminoC₁₋₈alkylenecarbonyl,        C₁₋₈alkoxyC₁₋₈alkylenecarbonyl, hydroxyC₁₋₈alkylenecarbonyl,        hydroxyC₁₋₈alkoxycarbonyl, aminocarbonyl, amino, C₁₋₈        alkoxycarbonylamino, arylC₁₋₈ alkoxycarbonylamino, hydroxyl,        C₁₋₈ alkoxy, C₁₋₈alkylsulfonyl, aminoC₁₋₈alkylenesulfonyl,        aminosulfonyl, C₁₋₈alkyleneaminoC₁₋₈alkylenesulfonyl,        C₁₋₈alkoxyC₁₋₈alkylenesulfonyl, hydroxyC₁₋₈alkylenesulfonyl,        hydroxyC₁₋₈alkoxysulfonyl, aminosulfonyl, oxo, halo, phenyl        heterocyclyl and C₁₋₈alkylheterocyclyl; and    -   (e) heterocyclyl; with from 1 to 2 substituents, R^(7e),        independently selected from the group consisting of C₁₋₈alkyl,        C₁₋₈alkoxy, halo, C₁₋₈alkylsulfonyl and heterocyclyl;

each E¹ is independently selected from the group consisting of C₁₋₈alkyl, C₂₋₈ alkenyl, C₂₋₈alkynyl, C₁₋₈alkoxy, C₁₋₈alkylthio,aminocarbonyl, C₁₋₈alkoxycarbonylC₁₋₈alkylene,C₁₋₈alkoxycarbonylC₁₋₈C₁₋₈alkoxy, C₁₋₈alkoxycarbonylamino, oxo, halo,cyano, haloC₁₋₈alkyl, haloC₁₋₈alkoxy, aminosulfonyl, heteroarylsulfinyl;amino, hydroxyl, C₁₋₈arylalkylene, phenyl, aminoC₁₋₈alkyl, aminoC₃₋₈cycloalkyl, heterocyclyl, heteraryl and heterocyclylC₁₋₈alkylene;

each R^(1a), R^(1b) and R^(1c) is independently selected from the groupconsisting of: H, C₁₋₈alkyl, hydroxyC₁₋₈ alkyl, amino, C₁₋₈alkylamino,C₁₋₈alkoxycarbonylaminoC₁₋₈ alkylene, C₃₋₈cycloalkyl, heteroaryl,C₁₋₈alkylC₃₋₈cycloalkyl, C₁₋₈alkylthioC₁₋₈ alkyl,C₁₋₈alkylsulfonylC₁₋₈alkylene, aminocarbonyl, aryl, and heterocyclyl;wherein the aryl is optionally substituted by hydroxyl, C₁₋₈alkoxy, haloor haloC₁₋₈alkyl; or taken together with R³ and the atoms to which theyare attached to form a C₃₋₈ cycloalkyl or heterocycloalkyl ring;

R² is selected from the group consisting of H, amino,arylC₁₋₈alkoxycarbonylamino and hydroxyl;

R³ is selected from the group consisting of H, C₁₋₈alkyl,C₁₋₈alkylamino, amino C₁₋₈alkylaminoC₁₋₈alkyl, C₁₋₈alkoxyC₁₋₈alkylene,hydroxyC₁₋₈alkyl and hydroxyC₁₋₈alkoxy; or may be combined with R^(1c)or R⁴ and the atoms to which they are attached to form a C₃₋₈cycloalkylor heterocyclyl ring;

R⁴ is H or alkyl or may be combined with R³ and the atoms to which theyare attached to form a C₃₋₈ cycloalkyl or heterocyclyl ring;

the subscript n is 0, 1, 2, 3 or 4; and

-   -   the subscript m is an integer of 1, 2 or 3; and the wavy line        indicates the point of attachment to the rest of the molecule.

The present invention provides in another embodiment, a compound whereinD¹ is C₃₋₈cycloalkyl.

The present invention provides in another embodiment, a compound whereinD¹ is selected from the group consisting of cyclopropyl, cyclobutyl,cyclopentyl and cyclohexyl.

The present invention provides in another embodiment, a compound whereinD¹ is heterocyclyl.

The present invention provides in another group of embodiments, acompound wherein any of the heterocyclyl groups of formula I is selectedfrom the group consisting of:

The present invention provides in another embodiment, a compound whereinthe heterocyclyl is selected from the group consisting of:

The present invention provides in another group of embodiments, acompound wherein a heteroaryl group of the compound of formula I isselected from the group consisting of:

each of which is optionally substituted with from 1 to 2 substituentsindependently selected from the group consisting of: C₁₋₈ alkyl, amino,hydroxyl, oxo, halo, C₁₋₈alkoxy, hydroxyC₁₋₈alkyl-, aminoC₁₋₈alkyl,C₁₋₈alkylcarbonyl, haloC₁₋₈alkyl, C₃₋₈cycloalkyl, C₁₋₈-aminocycloalkyl,aminoC₁₋₈alkylenecarbonyl, aminocarbonyl,C₁₋₈alkyleneaminoC₁₋₈alkylenecarbonyl, C₁₋₈alkoxyC₁₋₈alkylenecarbonyl,hydroxyC₁₋₈alkylenecarbonyl, hydroxyC₁₋₈alkoxycarbonyl,C₁₋₈alkoxycarbonylamino, aryl, arylC₁₋₈alkoxycarbonylamino,C₁₋₈alkylsulfonyl, aminoC₁₋₈alkylenesulfonyl, aminosulfonyl,C₁₋₈alkyleneaminoC₁₋₈alkylenesulfonyl, C₁₋₈alkoxyC₁₋₈alkylenesulfonyl,hydroxyC₁₋₈alkylenesulfonyl, hydroxyC₁₋₈alkoxysulfonyl, aminosulfonyl,and C₁₋₈alkylheterocyclyl.

The present invention provides in another group of embodiments, acompound wherein a heteroaryl group of the compound of formula I is apolycyclic heteroaryl group selected from the group consisting of:

optionally substituted with from 1 to 3 R^(7d) substituentsindependently selected from the group consisting of: C₁₋₈ alkyl,C₁₋₈alkylcarbonyl, C₁₋₈-aminocycloalkyl, aminoC₁₋₈alkylenecarbonyl,aminocarbonyl, C₁₋₈alkyleneaminoC₁₋₈alkylenecarbonyl,C₁₋₈alkoxyC₁₋₈alkylenecarbonyl, hydroxyC₁₋₈alkylenecarbonyl,hydroxyC₁₋₈alkoxycarbonyl, aminocarbonyl, amino, C₁₋₈alkoxycarbonylamino, aryl, arylC₁₋₈alkoxycarbonylamino, hydroxyl, C₁₋₈alkoxy, C₁₋₈alkylsulfonyl, aminoC₁₋₈alkylenesulfonyl, aminosulfonyl,C₁₋₈alkyleneaminoC₁₋₈alkylenesulfonyl, C₁₋₈alkoxyC₁₋₈alkylenesulfonyl,hydroxyC₁₋₈alkylenesulfonyl, hydroxyC₁₋₈alkoxysulfonyl, aminosulfonyl,oxo, halo, phenyl and C₁₋₈alkylheterocyclyl; and the wavy line indicatesthe point of attachment to the rest of the molecule.

The present invention provides in another embodiment, a compoundwherein: D¹ is bicyclic heteroaryl.

The present invention provides in another embodiment, a compound whereinD¹ is selected from the group consisting of:

optionally substituted with from 1 to 3 R^(1d) substituentsindependently selected from the group consisting of: C₁₋₈ alkyl,C₁₋₈alkylcarbonyl, aminoC₁₋₈alkylenecarbonyl, aminocarbonyl,C₁₋₈alkyleneaminoC₁₋₈alkylenecarbonyl, C₁₋₈alkoxyC₁₋₈alkylenecarbonyl,hydroxyC₁₋₈alkylenecarbonyl, hydroxyC₁₋₈alkoxycarbonyl, aminocarbonyl,amino, C₁₋₈ alkoxycarbonylamino, arylC₁₋₈ alkoxycarbonylamino, hydroxyl,C₁₋₈ alkoxy, C₁₋₈alkylsulfonyl, aminoC₁₋₈alkylenesulfonyl,aminosulfonyl, C₁₋₈ alkyleneaminoC₁₋₈alkylenesulfonyl,C₁₋₈alkoxyC₁₋₈alkylenesulfonyl, hydroxyC₁₋₈alkylenesulfonyl,hydroxyC₁₋₈alkoxysulfonyl, aminosulfonyl, oxo, halo, phenyl andC₁₋₈alkylheterocyclyl; and the wavy line indicates the point ofattachment to the rest of the molecule.

The present invention provides in another embodiment, a compound whereinD¹ is selected from the group consisting of:

and the wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another embodiment, a compound whereinD¹ is selected from the group consisting of:

and the wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another embodiment, a compound whereinD¹ is selected from the group consisting of:

and the wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another embodiment, a compound whereinR⁵ is selected from the group consisting of

-   -   each of which is optionally substituted with from 1 to 2        substituents independently selected from the group consisting of        C₁₋₈alkyl, hydroxyC₁₋₈alkyl-, aminoC₁₋₈alkyl, C₁₋₈alkylcarbonyl,        aminocarbonyl, hydroxy, oxo, halo, haloC₁₋₈alkyl, aminosulfonyl        and C₃₋₈cycloalkyl.

The present invention provides in another embodiment, a compound whereinR⁵ is selected from the group consisting of

each of which is optionally substituted with from 1 to 2 substituentsindependently selected from the group consisting of C₁₋₈alkyl,hydroxyC₁₋₈alkyl-, aminoC₁₋₈alkyl, C₁₋₈alkylcarbonyl, aminocarbonyl,hydroxy, oxo, halo, haloC₁₋₈alkyl, aminosulfonyl and C₃₋₈cycloalkyl.

The present invention provides in another embodiment, a compound whereinR⁵ is selected from the group consisting of

The present invention provides in another embodiment, a compound whereineach E¹ is independently selected from the group consisting ofC₁₋₈alkyl, heteroaryl, heterocyclyl, halo, C₁₋₈haloalkyl, C₁₋₈alkoxy,C₁₋₈acyl, aminoC₁₋₈alkyl, aminosulfonyl, C₁₋₈alkylsulfonyl andacylamino.

The present invention provides in another embodiment, a compound whereineach E¹ is independently selected from the group consisting of

The present invention provides in another embodiment, a compound whereineach E¹ is independently selected from the group consisting of

The present invention provides in another embodiment, a compound whereinR² is amino.

The present invention provides in another embodiment, a compound whereinthe moiety:

-   -   wherein X and Y is each independently selected from the group        consisting of: CH₂, NH, NCOCH₃ and S.

The present invention provides in another embodiment, a compound whereinthe moiety:

is selected from the group consisting of:

wherein each R^(8a) and R^(8b) is independently H, hydroxyl, halo or ifon adjacent carbon atoms, may be combined with the atoms to which theyare attached to form a fused benzene ring; and

-   -   each R^(9a) and R^(9b) is independently H, hydroxyl, halo or, if        on adjacent carbon atoms, may be combined with the atoms to        which they are attached to form a fused benzene ring; and the        wavy line indicates the point of attachment to the rest of the        molecule.

The present invention provides in another embodiment, a compound whereinthe moiety:

wherein each R^(8a) and R^(8b) is independently H or may be combinedwith the atoms to which they are attached to form a fused benzene ring;and each R^(9a) and R^(9b) is independently H or may be combined withthe atoms to which they are attached to form a fused benzene ring; andthe wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another embodiment, a compoundwherein: the moiety:

-   -   wherein each R^(8a) and R^(8b) is independently H or may be        combined with the atoms to which they are attached to form a        fused benzene ring; and the wavy line indicates the point of        attachment to the rest of the molecule.

The present invention provides in another group of embodiments, acompound wherein the moiety:

The present invention provides in another group of embodiments, acompound wherein the moiety:

The present invention provides in another group of embodiments, acompound wherein R^(1c) is selected from the group consisting of:C₁₋₈alkyl, hydroxyC₁₋₈alkyl, C₁₋₈alkoxyC₁₋₈alkyl and haloC₁₋₈alkyl;

R² is selected from the group consisting of H, amino and C₁₋₈alkylamino;

R³ is selected from the group consisting of H, C₁₋₈alkyl,aminoC₁₋₈alkyl, carboxy, aminoC₁₋₈alkylaminoC₁₋₈alkyl,C₁₋₈alkoxyC₁₋₈alkyl, hydroxyC₁₋₈alkyl, carboxyC₁₋₈alkyl,C₃₋₈cycloalkylC₁₋₈alkyl, aryloxyC₁₋₈alkyl, arylC₁₋₈alkyl,heteroarylC₁₋₈alkyl, and hydroxyC₁₋₈alkoxy; or may be combined withR^(1c) or R⁴ and the atoms to which they are attached to form aC₃₋₈cycloalkyl or heterocyclyl ring;

R⁴ is H or alkyl or may be combined with R³ and the atoms to which theyare attached to form a C₃₋₈ cycloalkyl or heterocyclyl ring.

The present invention provides in another group of embodiments, acompound having the formula:

-   -   wherein each X and Y is independently selected from the group        consisting of: CH₂, NH, NCOCH₃ and S.

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

-   -   wherein each X and Y is independently selected from the group        consisting of: CH₂, NH, NCOCH₃ and S.

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having a formula selected from the group consisting of:

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

-   -   wherein each X and Y is independently selected from the group        consisting of: CH₂, NH, NCOCH₃ and S.

The present invention provides in another embodiment, a compoundselected from the group consisting of:

The present invention provides in another embodiment, a compoundselected from the group consisting of:

The present invention provides in another embodiment, a compound, havingthe formula:

The present invention provides in another embodiment, a compound, havingthe formula:

The present invention provides in another embodiment, a compound, havingthe formula:

The present invention provides in another embodiment, a compoundselected from the group consisting of:

The present invention provides in another embodiment, a compound whereinthe moiety:

and the wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having a formula selected from the group consisting of:

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another embodiment, a compound whereinR^(1a) is selected from the group consisting of C₁₋₈alkyl,hydroxylC₁₋₈alkyl, C₃₋₈cycloalkyl, aryl, heteroaryl and heterocyclyl.

The present invention provides in another embodiment, a compound whereinR^(1a) is selected from the group consisting of C₁₋₈alkyl,C₃₋₈cycloalkyl, aryl, heteroaryl and heterocyclyl.

The present invention provides in another group of embodiments, acompound wherein the moiety:

is selected from the group consisting of:

The present invention provides in another embodiment, a compound havingthe structure selected from the group consisting of formula IIa-c:

-   -   or a pharmaceutically acceptable salt thereof, wherein:    -   E^(2a) is selected from the group consisting of C₁₋₈ alkoxy,        C₁₋₈alkylC₃₋₈cycloalkylcarbonylamino,        C₁₋₈alkoxyC₁₋₈alkylenecarbonylamino-, C₁₋₈alkoxycarbonylamino        and C₁₋₈alkylcarbonylamino;    -   R³ is selected from the group consisting of H, halo, C₁₋₈alkyl        and C₁₋₈alkoxy; and    -   R⁴ is selected from the group consisting of H, C₁₋₈alkyl and        C₁₋₈alkoxy C₁₋₈alkylene.

The present invention provides in another embodiment, a compound havingformula (IId):

or a pharmaceutically acceptable salt thereof,wherein R³ is H or C₁₋₈alkyl.

The present invention provides in another embodiment, a compound havingformula:

The present invention provides in another embodiment, a compound havingformula (IIe):

-   -   wherein: D² is a bicyclic aryl group.

The present invention provides in another embodiment, a compoundwherein: D² is naphthyl, optionally substituted with from 1 to 2substituents, E^(2b), independently selected from the group consistingof halo, C₁₋₈alkoxy and C₁₋₈alkylaminocarbonyl.

The present invention provides in another embodiment, a compound whereinD² is

The present invention provides in another embodiment, a compound whereinD² is

The present invention provides in another embodiment, a compound whereinD² is selected from the group consisting of:

and the wavy line indicates the point of attachment to the rest of themolecule.

The present invention provides in another embodiment, a compound, havingthe formula

The present invention provides in another embodiment, a compound havingthe structure found in the Examples.

The present invention provides in another embodiment, a compound havingthe structure found in the tables.

It is understood that in another group of embodiments, any of the aboveembodiments may also be combined with other embodiments listed herein,to form other embodiments of the invention.

b. Methods of Synthesis

The compounds of the present invention may be prepared by known organicsynthesis techniques, including the methods described in more detail inthe Examples. In general, the compounds of structure (I) above may bemade by the following FIG. 3, wherein all substituents are as definedabove unless indicated otherwise.

Compounds having formula I may be prepared according to FIG. 3.Carboxylic acid 1.1 is converted to acid chloride 1.2 via a one-stepprocedure by treatment with a chlorination agent, such as thionylchloride, and esterification with an alcohol, such as ethanol, to formcompound 1.3 using conditions similar to that described below. Ester 1.3is dichlorinated with a chlorinating agent, such as phosphorousoxychloride. Selective displacement of the 4-chloro group of the2,4-dichloropyrimidine by an appropriate amine, such as E¹-D¹-NH₂(available commercially or synthesized using methods known to thoseskilled in the art), under basic conditions, such as withdiisopropylamine (DIA), provides compounds of formula 1.5. Subsequenthydrolysis of the ester, displacement of the second chloro group withEDC and treatment with ammonia gives compound 1.7. Benzotriazolyl ethercompound 1.7 may also be prepared through a linear route. Displacementof the benzotriazolyl ether group with an appropriate amine, such asY¹—NH₂ (available commercially or synthesized using methods known tothose skilled in the art), gives the desired product I, wherein E¹-D¹and Y¹ are as previously defined.

One skilled in the art will recognize that in certain embodiments ofstructure (I) when E¹-D¹ or Y¹ comprises a terminal heteroatom, it maybe advantageous to use a protecting group strategy. The protecting groupcan be removed using methods known to those skilled in the art to yieldcompounds of structure (1).

The compounds of the present invention may generally be utilized as thefree base. Alternatively, the compounds of this invention may be used inthe form of acid addition salts as described below.

c. Inhibition of syk Kinase

The activity of a specified compound as an inhibitor of a syk kinase maybe assessed in vitro or in vivo. In some embodiments, the activity of aspecified compound can be tested in a cellular assay. Selectivity couldalso be ascertained in biochemical assays with isolated kinases.

Similar types of assays can be used to assess JAK kinase inhibitoryactivity and to determine the degree of selectivity of the particularcompound as compared to syk kinase. One means of assaying for suchinhibition is detection of the effect of the compounds of the presentinvention on the upregulation of downstream gene products. In theRamos/IL4 assay, B-cells are stimulated with the cytokine Interleukin-4(IL-4) leading to the activation of the JAK/Stat pathway throughphosphorylation of the JAK family kinases, JAK1 and JAK3, which in turnphosphorylate and activate the transcription factor Stat-6. One of thegenes upregulated by activated Stat-6 is the low affinity IgE receptor,CD23. To study the effect of inhibitors (e.g., the 2,4-substitutedpyrimindinediamine compounds described herein) on the JAK1 and JAK3kinases, human Ramos B-cells are stimulated with human IL-4. 10′post-stimulation, cells are subjected to intracellular flow cytometry tomeasure the extent of STAT-6 phosphorylation. 20 to 24 hourspost-stimulation, cells are stained for upregulation of CD23 andanalyzed using flow cytometry. A reduction of the amount ofphosphohorylated STAT-6 and/or cell surface CD23 present compared tocontrol conditions indicates that the test compound actively inhibitsthe JAK kinase pathway.

Additionally, IL-6 stimulation of Ramos B-cells induces JAKs 1, 2, andTyk2, leading to Stat-3 and Erk phosphorylation. 10′ post-stimulation,cells are subjected to intracellular flow cytometry to measure theability of compound to inhibit these phosphorylation events. Tospecifically measure the activity of JAK2, the CellSensor irfl-bla HELcell line expressing the beta-lactamase reporter gene controlled byStat5 will be used (Invitrogen, Carlsbad, Calif.). These cells express aconstituitively active JAK2 mutant (JAK2V617F), found naturally inmyeloproliferative neoplasms (Constantinescu, S., et. al, TrendsBiochem. Sci., 2008; 33:122-31). A reduction in the amount ofbeta-lactamase reporter gene expression is used a measure of the JAK2inhibitory activity of compounds.

The activity of the compounds of the invention may additionally becharacterized by assaying the effect of the compounds of the presentinvention described herein on A549 lung epithelial cells and U937 cells.A549 lung epithelial cells and U937 cells up-regulate ICAM-1 (CD54)surface expression in response to a variety of different stimuli.Therefore, using ICAM-1 expression as readout, test compound effects ondifferent signaling pathways can be assessed in the same cell type.Stimulation with IL-1β through the IL-1 β receptor activates theTRAF6/NFκB pathway resulting in up-regulation of ICAM-1. IFN.gamma.induces ICAM-1 up-regulation through activation of the JAK1/JAK2pathway. The up-regulation of ICAM-1 can be quantified by flow cytometryacross a compound dose curve and EC₅₀ values are calculated.

The activity of the compounds of the invention may additionally becharacterized by assaying the effect of the compounds of the presentinvention described herein on A549 lung epithelial cells and U937 cells.A549 lung epithelial cells and U937 cells up-regulate ICAM-1 (CD54)surface expression in response to a variety of different stimuli.Therefore, using ICAM-1 expression as readout, test compound effects ondifferent signaling pathways can be assessed in the same cell type.Stimulation with IL-1β through the IL-1 β receptor activates theTRAF6/NFκB pathway resulting in up-regulation of ICAM-1. IFN.gamma.induces ICAM-1 up-regulation through activation of the JAK1/JAK2pathway. The up-regulation of ICAM-1 can be quantified by flow cytometryacross a compound dose curve and EC₅₀ values are calculated. Exemplaryassays of this type are described in greater detail in the Examples.

Active compounds as described herein generally inhibit the JAK kinasepathway with an IC₅₀ in the range of about 1 mM or less, as measured inthe assays described herein. Of course, skilled artisans will appreciatethat compounds which exhibit lower IC₅₀s, (on the order, for example, of100 μM, 75 μM, 50 μM, 40 μM, 30 μM, 20 μM, 15 μM, 10 μM, 5 μM, 1 μM, 500nM, 100 nM, 10 nM, 1 nM, or even lower) can be particularly useful intherapeutic applications. In instances where activity specific to aparticular cell type is desired, the compound can be assayed foractivity with the desired cell type and counter-screened for a lack ofactivity against other cell types. The desired degree of “inactivity” insuch counter screens, or the desired ratio of activity vs. inactivity,may vary for different situations and can be selected by the user.

The active compounds also typically inhibit IL-4 stimulated expressionof CD23 in B-cells with an IC₅₀ in the range of about 20 μM or less,typically in the range of about 10 μM, 1 μM, 500 nM, 100 nM, 10 nM, 1nM, or even lower. A suitable assay that can be used is the assaydescribed in the Examples, “Assay for Ramos B-cell Line Stimulated withIL-4.” In certain embodiments, the active compounds of the presentinvention have an IC₅₀ of less than or equal to 5 μM, greater than 5 μMbut less than 20 μM, greater than 20 μM, or greater than 20 μM but lessthan 50 μM in the assay described in the Examples.

The active compounds also typically inhibit expression of ICAMI (CD54)induced by IFN.gamma. exposure in U937 or A549 cells with an IC₅₀ in therange of about 20 μM or less, typically in the range of about 10 μM, 1μM, 500 nM, 100 nM, 10 nM, 1 nM, or even lower. The IC₅₀ againstexpression of ICAM (CD54) in IFN.gamma. stimulated cells can bedetermined in a functional cellular assay with an isolated A549 or U937cell line. Suitable assays that can be used are the assays described inthe Examples, “A549 Epithelial Line Stimulated with IFNγ” and “U937IFN.gamma. ICAM1 FACS Assay,” respectively. In certain embodiments, theactive compounds of the present invention have an IC₅₀ of less than orequal to 20 μM, greater than 20 μM, or greater than 20 μM but less than50 μM in the assays described in the Examples.

d. Compositions and Methods of Administration

The present invention further provides compositions comprising one ormore compounds of formula (I) or a pharmaceutically acceptable salt,ester or prodrug thereof, and a pharmaceutically acceptable carrier ordiluent. It will be appreciated that the compounds of formula (I)) inthis invention may be derivatized at functional groups to provideprodrug derivatives which are capable of conversion back to the parentcompounds in vivo. Examples of such prodrugs include the physiologicallyacceptable and metabolically labile ester derivatives, such asmethoxymethyl esters, methylthiomethyl esters, or pivaloyloxymethylesters derived from a hydroxyl group of the compound or a carbamoylmoiety derived from an amino group of the compound. Additionally, anyphysiologically acceptable equivalents of the compounds of formula (I),similar to metabolically labile esters or carbamates, which are capableof producing the parent compounds of formula (I) in vivo, are within thescope of this invention.

As used herein, the term “pharmaceutically acceptable salts” refers toany acid or base addition salt whose counter-ions are non-toxic to thepatient in pharmaceutical doses of the salts. A host of pharmaceuticallyacceptable salts are well known in the pharmaceutical field. Ifpharmaceutically acceptable salts of the compounds of this invention areutilized in these compositions, those salts are preferably derived frominorganic or organic acids and bases. Included among such acid salts arethe following: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphor sulfonate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,fumarate, lucoheptanoate, glycerophosphate, hemisulfate, heptanoate,hexanoate, hydrochloride, hydrobromide, hydroiodide,2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate,2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectinate,persulfate, 3-phenyl-propionate, picrate, pivalate, propionate,succinate, tartrate, thiocyanate, tosylate, undecanoate, hydrohalides(e.g., hydrochlorides and hydrobromides), sulphates, phosphates,nitrates, sulphamates, malonates, salicylates,methylene-bis-b-hydroxynaphthoates, gentisates, isethionates,di-p-toluoyltartrates, ethanesulphonates, cyclohexylsulphamates,quinates, and the like. Pharmaceutically acceptable base addition saltsinclude, without limitation, those derived from alkali or alkaline earthmetal bases or conventional organic bases, such as triethylamine,pyridine, piperidine, morpholine, N-methylmorpholine, ammonium salts,alkali metal salts, such as sodium and potassium salts, alkaline earthmetal salts, such as calcium and magnesium salts, salts with organicbases, such as dicyclohexylamine salts, N-methyl-D-glucamine, and saltswith amino acids such as arginine, lysine, and so forth.

Furthermore, the basic nitrogen-containing groups may be quaternizedwith agents like lower alkyl halides, such as methyl, ethyl, propyl andbutyl chlorides, bromides and iodides; dialkyl sulfates, such asdimethyl, diethyl, dibutyl and diamyl sulfates, long chain halides, suchas decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides;aralkyl halides, such as benzyl and phenethyl bromides and others. Wateror oil-soluble or dispersible products are thereby obtained.

The compounds utilized in the compositions and methods of this inventionmay also be modified by appending appropriate functionalities to enhanceselective biological properties. Such modifications are known in the artand include those which increase biological penetration into a givenbiological system (e.g., blood, lymphatic system, central nervoussystem, etc.), increase oral availability, increase solubility to allowadministration by injection, alter metabolism and alter rate ofexcretion.

The pharmaceutical compositions of the invention can be manufactured bymethods well known in the art such as conventional granulating, mixing,dissolving, encapsulating, lyophilizing, or emulsifying processes, amongothers. Compositions may be produced in various forms, includinggranules, precipitates, or particulates, powders, including freezedried, rotary dried or spray dried powders, amorphous powders, tablets,capsules, syrup, suppositories, injections, emulsions, elixirs,suspensions or solutions. Formulations may optionally containstabilizers, pH modifiers, surfactants, bioavailability modifiers andcombinations of these.

The term “unit dosage form” refers to physically discrete units suitableas unitary dosages for human subjects and other mammals, each unitcontaining a predetermined quantity of drug calculated to produce thedesired onset, tolerability, and/or therapeutic effects, in associationwith a suitable pharmaceutical excipient (e.g., an ampoule). Inaddition, more concentrated compositions may be prepared, from which themore dilute unit dosage compositions may then be produced. The moreconcentrated compositions thus will contain substantially more than,e.g., at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more times the amountof one or more syk inhibitors.

Methods for preparing such dosage forms are known to those skilled inthe art (see, for example, R EMINGTON'S PHARMACEUTICAL SCIENCES, 18THED., Mack Publishing Co., Easton, Pa. (1990)). In addition,pharmaceutically acceptable salts of the syk inhibitors of the presentinvention (e.g., acid addition salts) may be prepared and included inthe compositions using standard procedures known to those skilled in theart of synthetic organic chemistry and described, e.g., by J. March,Advanced Organic Chemistry: Reactions, Mechanisms and Structure, 4^(th)Ed. (New York: Wiley-Interscience, 1992).

The compositions typically include a conventional pharmaceutical carrieror excipient and may additionally include other medicinal agents,carriers, adjuvants, diluents, tissue permeation enhancers,solubilizers, and the like. Preferably, the composition will containabout 0.01% to about 90%, preferably about 0.1% to about 75%, morepreferably about 0.1% to 50%, still more preferably about 0.1% to 10% byweight of one or more syk inhibitors, with the remainder consisting ofsuitable pharmaceutical carrier and/or excipients. Appropriateexcipients can be tailored to the particular composition and route ofadministration by methods well known in the art, e.g., R EMINGTON'SPHARMACEUTICAL SCIENCES, supra.

Pharmaceutically acceptable carriers that may be used in thesecompositions include ion exchangers, alumina, aluminum stearate,lecithin, serum proteins, such as human serum albumin, buffersubstances, such as phosphates, glycine, sorbic acid, potassium sorbate,partial glyceride mixtures of saturated vegetable fatty acids, water,salts or electrolytes, such as protamine sulfate, disodium hydrogenphosphate, potassium hydrogen phosphate, sodium chloride, zinc salts,colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone,cellulose-based substances, polyethylene glycol, sodiumcarboxymethylcellulose, polyacrylates, waxes,polyethylene-polyoxypropylene-block polymers, polyethylene glycol andwool fat.

Examples of suitable excipients include, but are not limited to,lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia,calcium phosphate, alginates, tragacanth, gelatin, calcium silicate,microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water,saline, syrup, methylcellulose, ethylcellulose,hydroxypropylmethylcellulose, and polyacrylic acids such as Carbopols.The compositions can additionally include lubricating agents such astalc, magnesium stearate, and mineral oil; wetting agents; emulsifyingagents; suspending agents; preserving agents such as methyl-, ethyl-,and propyl-hydroxy-benzoates; pH adjusting agents such as inorganic andorganic acids and bases; sweetening agents; and flavoring agents.

Administration of a composition comprising one or more syk inhibitorswith one or more suitable pharmaceutical excipients as advantageous canbe carried out via any of the accepted modes of administration. Thus,administration can be, for example, oral, topical, intravenous,subcutaneous, transcutaneous, transdermal, intramuscular, intra-joint,parenteral, intra-arteriole, intradermal, intraventricular,intracranial, intraperitoneal, intralesional, intranasal, rectal,vaginal, by inhalation or via an implanted reservoir. The term“parenteral” as used herein includes subcutaneous, intravenous,intramuscular, intra-articular, intra-synovial, intrasternal,intrathecal, intrahepatic, intralesional and intracranial injection orinfusion techniques. Preferably, the compositions are administeredorally or intravenously. The formulations of the invention may bedesigned as short-acting, fast-releasing, or long-acting. Still further,compounds can be administered in a local rather than systemic means,such as administration (e.g., injection) as a sustained releaseformulation. According to a representative embodiment, the compositionsof this invention are formulated for pharmaceutical administration to amammal, preferably a human being.

The compositions of the present invention containing one or more sykinhibitors can be administered repeatedly, e.g., at least 2, 3, 4, 5, 6,7, 8, or more times, or the composition may be administered bycontinuous infusion. Suitable sites of administration include, but arenot limited to, skin, bronchial, gastrointestinal, anal, vaginal, eye,and ear. The formulations may take the form of solid, semi-solid,lyophilized powder, or liquid dosage forms, such as, for example,tablets, pills, capsules, powders, solutions, suspensions, emulsions,suppositories, retention enemas, creams, ointments, lotions, gels,aerosols, or the like, preferably in unit dosage forms suitable forsimple administration of precise dosages.

The pharmaceutical compositions of this invention may be in any orallyacceptable dosage form, including tablets, capsules, cachets, emulsions,suspensions, solutions, syrups, elixirs, sprays, boluses, lozenges,powders, granules, and sustained-release formulations. Suitableexcipients for oral administration include pharmaceutical grades ofmannitol, lactose, starch, magnesium stearate, sodium saccharine,talcum, cellulose, glucose, gelatin, sucrose, magnesium carbonate, andthe like. In the case of tablets for oral use, carriers that arecommonly used include lactose and corn starch. Lubricating agents, suchas magnesium stearate, are also typically added. For a capsule form,useful diluents include lactose and dried cornstarch. When aqueoussuspensions are required for oral use, the active ingredient is combinedwith emulsifying and suspending agents. If desired, certain sweetening,flavoring or coloring agents may also be added.

In some embodiments, the compositions take the form of a pill, tablet,or capsule, and thus, the composition can contain, along with one ormore syk inhibitors, a diluent such as lactose, sucrose, dicalciumphosphate, and the like; a disintegrant such as starch or derivativesthereof; a lubricant such as magnesium stearate and the like; and/or abinder such a starch, gum acacia, polyvinylpyrrolidone, gelatin,cellulose and derivatives thereof. A tablet can be made by anycompression or molding process known to those of skill in the art.Compressed tablets may be prepared by compressing in a suitable machinethe syk inhibitors in a free-flowing form, e.g., a powder or granules,optionally mixed with accessory ingredients, e.g., binders, lubricants,diluents, disintegrants, or dispersing agents. Molded tablets can bemade by molding in a suitable machine a mixture of the powdered sykinhibitors with any suitable carrier.

Alternatively, the pharmaceutical compositions of this invention may bein the form of suppositories for rectal administration. These may beprepared by mixing the agent with a suitable non-irritating excipientwhich is solid at room temperature but liquid at rectal temperature andtherefore will melt in the rectum to release the drug. Such materialsinclude cocoa butter, beeswax, polyethylene glycol (PEG), hard fat,and/or hydrogenated cocoglyceride. Compositions suitable for rectaladministration may also comprise a rectal enema unit containing one ormore syk inhibitors and pharmaceutically-acceptable vehicles (e.g., 50%aqueous ethanol or an aqueous salt solution) that are physiologicallycompatible with the rectum and/or colon. The rectal enema unit containsan applicator tip protected by an inert cover, preferably comprised ofpolyethylene, lubricated with a lubricant such as white petrolatum, andpreferably protected by a one-way valve to prevent back-flow of thedispensed formula. The rectal enema unit is also of sufficient length,preferably two inches, to be inserted into the colon via the anus.

Liquid compositions can be prepared by dissolving or dispersing one ormore syk inhibitors and optionally one or more pharmaceuticallyacceptable adjuvants in a carrier such as, for example, aqueous saline,aqueous dextrose, glycerol, ethanol, and the like, to form a solution orsuspension, e.g., for oral, topical, or intravenous administration.Pharmaceutical formulations may be prepared as liquid suspensions orsolutions using a sterile liquid, such as oil, water, alcohol, andcombinations thereof. Pharmaceutically suitable surfactants, suspendingagents or emulsifying agents, may be added for oral or parenteraladministration. Suspensions may include oils, such as peanut oil, sesameoil, cottonseed oil, corn oil and olive oil. Suspension preparation mayalso contain esters of fatty acids, such as ethyl oleate, isopropylmyristate, fatty acid glycerides and acetylated fatty acid glycerides.Suspension formulations may include alcohols, such as ethanol, isopropylalcohol, hexadecyl alcohol, glycerol and propylene glycol. Ethers, suchas poly(ethyleneglycol), petroleum hydrocarbons, such as mineral oil andpetrolatum, and water may also be used in suspension formulations.

The pharmaceutical compositions of this invention may also be in atopical form, especially when the target of treatment includes areas ororgans readily accessible by topical application, including diseases ofthe eye, the skin, or the lower intestinal tract. Suitable topicalformulations are readily prepared for each of these areas or organs. Fortopical administration, the composition containing one or more sykinhibitors can be in the form of emulsions, lotions, gels, foams,creams, jellies, solutions, suspensions, ointments, and transdermalpatches.

Topical application for the lower intestinal tract may be effected in arectal suppository formulation (see above) or in a suitable enemaformulation. Topically-transdermal patches may also be used. For topicalapplications, the pharmaceutical compositions may be formulated in asuitable ointment containing the active component suspended or dissolvedin one or more carriers. Carriers for topical administration of thecompounds of this invention include, but are not limited to, mineraloil, liquid petrolatum, white petrolatum, propylene glycol,polyoxyethylene, polyoxypropylene compound, emulsifying wax and water.Alternatively, the pharmaceutical compositions may be formulated in asuitable lotion or cream containing the active components suspended ordissolved in one or more pharmaceutically acceptable carriers. Suitablecarriers include mineral oil, sorbitan monostearate, polysorbate 60,cetyl esters, wax, cetyl alcohol, 2-octyldodecanol, benzyl alcohol andwater.

The pharmaceutical compositions of this invention may also beadministered by nasal aerosol or inhalation. For delivery by inhalation,the compositions can be delivered as a dry powder or in liquid form viaa nebulizer. Such compositions are prepared according to techniquesknown in the art of pharmaceutical formulation and may be prepared assolutions in saline, employing benzyl alcohol or other suitablepreservatives, absorption promoters to enhance bioavailability,fluorocarbons and/or other conventional solubilizing or dispersingagents.

For ophthalmic use, the pharmaceutical compositions may be formulated asmicronized suspensions in isotonic, pH adjusted sterile saline, or,preferably, as solutions in isotonic, pH adjusted sterile saline, eitherwith our without a preservative, such as benzylalkonium chloride.Alternatively, for ophthalmic uses, the pharmaceutical compositions maybe formulated in an ointment, such as petrolatum.

For parenteral administration, the compositions can be in the form ofsterile injectable solutions and sterile packaged powders. Preferably,injectable solutions are formulated at a pH of about 4.5 to about 7.5.

Sterile injectable forms of the compositions of this invention may beaqueous or oleaginous suspension. These suspensions may be formulatedaccording to techniques known in the art using suitable dispersing orwetting agents and suspending agents. The sterile injectable preparationmay also be a sterile injectable solution or suspension in a non-toxicparenterally acceptable diluent or solvent, for example as a solution in1,3-butanediol. Among the acceptable vehicles and solvents that may beemployed are water, Ringer's solution and isotonic sodium chloridesolution. In addition, sterile, fixed oils are conventionally employedas a solvent or suspending medium. For this purpose, any bland fixed oilmay be employed including synthetic mono- or di-glycerides. Fatty acids,such as oleic acid and its glyceride derivatives are useful in thepreparation of injectables, as are natural pharmaceutically-acceptableoils, such as olive oil or castor oil, especially in theirpolyoxyethylated versions. These oil solutions or suspensions may alsocontain a long-chain alcohol diluent or dispersant, such ascarboxymethyl cellulose or similar dispersing agents which are commonlyused in the formulation of pharmaceutically acceptable dosage formsincluding emulsions and suspensions. Other commonly used surfactants,such as Tweens, Spans and other emulsifying agents or bioavailabilityenhancers which are commonly used in the manufacture of pharmaceuticallyacceptable solid, liquid, or other dosage forms may also be used for thepurposes of formulation. Compounds may be formulated for parenteraladministration by injection such as by bolus injection or continuousinfusion. A unit dosage form for injection may be in ampoules or inmulti-dose containers.

The compositions of the present invention can also be provided in alyophilized form. Such compositions may include a buffer, e.g.,bicarbonate, for reconstitution prior to administration, or the buffermay be included in the lyophilized composition for reconstitution with,e.g., water. The lyophilized composition may further comprise a suitablevasoconstrictor, e.g., epinephrine. The lyophilized composition can beprovided in a syringe, optionally packaged in combination with thebuffer for reconstitution, such that the reconstituted composition canbe immediately administered to a patient.

Any of the above dosage forms containing effective amounts are withinthe bounds of routine experimentation and within the scope of theinvention. A therapeutically effective dose may vary depending upon theroute of administration and dosage form. The representative compound orcompounds of the invention is a formulation that exhibits a hightherapeutic index. The therapeutic index is the dose ratio between toxicand therapeutic effects which can be expressed as the ratio between LD₅₀and ED₅₀. The LD₅₀ is the dose lethal to 50% of the population and theED₅₀ is the dose therapeutically effective in 50% of the population. TheLD₅₀ and ED₅₀ are determined by standard pharmaceutical procedures inanimal cell cultures or experimental animals.

Besides those representative dosage forms described above,pharmaceutically acceptable excipients and carriers and dosage forms aregenerally known to those skilled in the art and are included in theinvention. It should be understood that a specific dosage and treatmentregimen for any particular patient will depend upon a variety offactors, including the activity of the specific compound employed, theage, body weight, general health, sex and diet of the patient, and thetime of administration, rate of excretion, drug combination, judgment ofthe treating physician and severity of the particular disease beingtreated. The amount of active ingredient(s) will also depend upon theparticular compound and other therapeutic agent, if present, in thecomposition.

e. Methods of Use

The invention provides methods of inhibiting or decreasing syk activityas well as treating or ameliorating a syk associated state, symptom,condition, disorder or disease in a patient in need thereof (e.g., humanor non-human). In one embodiment, the syk associated state, symptom,condition, disorder or disease is mediated, at least in part by sykkinase activity. In more specific embodiments, the present inventionprovides a method for treating a condition or disorder mediated at leastin part by syk kinase activity is cardiovascular disease, inflammatorydisease or autoimmune disease.

In one embodiment, the invention provides methods for preventing ortreating a condition in a mammal characterized by undesired thrombosiscomprising the step of administering to the mammal a therapeuticallyeffective amount of a compound of the present invention. Such conditionsinclude, but are not limited, to restenosis, acute coronary syndrome,myocardial infarction, unstable angina, refractory angina, occlusivecoronary thrombosis occurring post-thrombolytic therapy or post-coronaryangioplasty, a thrombotically mediated cerebrovascular syndrome, embolicstroke, thrombotic stroke, transient ischemic attacks, venousthrombosis, deep venous thrombosis, pulmonary embolism, coagulopathy,disseminated intravascular coagulation, thrombotic thrombocytopenicpurpura, thromboangiitis obliterans, thrombotic disease associated withheparin-induced thrombocytopenia, thrombotic complications associatedwith extracorporeal circulation, thrombotic complications associatedwith instrumentation such as cardiac or other intravascularcatheterization, intra-aortic balloon pump, coronary stent or cardiacvalve, conditions requiring the fitting of prosthetic devices, and thelike.

In a further embodiment, the present invention provides a method fortreating thrombosis, immune thrombocytic purura, heparin inducedthrombocytopenia, dilated cardiomypathy, sickle cell disease,atherosclerosis, myocardial infarction, vacular inflammation, unstableangina or acute coronary syndromes.

In another embodiment, the present invention also provides a method fortreating allergy, asthma, theumatoid arthritis, B Cell mediated diseasesuch as Non-Hodgkin's Lymphoma, anti phospholipids syndrome, lupus,psoriasis, multiple sclerosis, end stage renal disease or chroniclymphocytic leukemia.

In another embodiment, the present invention provides a method fortreating hemolytic anemia or immune thrombocytopenic purpura.

The compounds described herein are also potent and/or selectiveinhibitors of JAK kinases. As a consequence of this activity, thecompounds can be used in a variety of in vitro, in vivo, and ex vivocontexts to regulate or inhibit JAK kinase activity, signaling cascadesin which JAK kinases play a role, and the biological responses effectedby such signaling cascades. For example, in one embodiment, thecompounds can be used to inhibit JAK kinase, either in vitro or in vivo,in virtually any cell type expressing the JAK kinase, such as inhematopoietic cells in which, for example, JAK3 is predominantlyexpressed. They may also be used to regulate signal transductioncascades in which JAK kinases, particularly JAK3, play a role. SuchJAK-dependent signal transduction cascades include, but are not limitedto, the signaling cascades of cytokine receptors that involve the commongamma chain, such as, for example, the IL-4, IL-7, IL-5, IL-9, IL-15 andIL-21, or IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 receptor signalingcascades. The compounds may also be used in vitro or in vivo toregulate, and in particular to inhibit, cellular or biological responsesaffected by such JAK-dependent signal transduction cascades. Suchcellular or biological responses include, but are not limited to,IL-4/ramos CD23 upregulation and IL-2 mediated T-cell proliferation.Importantly, the compounds can be used to inhibit JAK kinases in vivo asa therapeutic approach towards the treatment or prevention of diseasesmediated, either wholly or in part, by a JAK kinase activity (referredto herein as “JAK kinase mediated diseases”). Non-limiting examples ofJAK kinase mediated diseases that can be treated or prevented with thecompounds include, but are not limited to, the following: allergies;asthma; autoimmune diseases such as transplant rejection (e.g., kidney,heart, lung, liver, pancreas, skin, small intestine, large intestine,host versus graft reaction (HVGR), and graft versus host reaction(GVHR)), rheumatoid arthritis, and amyotrophic lateral sclerosis; T-cellmediated autoimmune diseases such as multiple sclerosis, psoraiasis, andSjogren's syndrome; Type II inflammatory diseases such as vascularinflammation (including vasculitis, arteritis, atherosclerosis, andcoronary artery disease); diseases of the central nervous system such asstroke; pulmonary diseases such as bronchitis obliteraus and primarypulmonary hypertension; solid, delayed Type IV hypersensitivityreactions; and hematologic malignancies such as leukemia and lymphomas.

Examples of diseases that are mediated, at least in part, by JAK kinasesthat can be treated or prevented according to the methods include, butare not limited to, allergies, asthma, autoimmune diseases such astransplant rejection (e.g., kidney, heart, lung, liver, pancreas, skin,host versus graft reaction (HVGR), etc.), rheumatoid arthritis, andamyotrophic lateral sclerosis, multiple sclerosis, psoraiasis andSjogren's syndrome, Type II inflammatory disease such as vascularinflammation (including vasculitis, ateritis, atherosclerosis andcoronary artery disease) or other inflammatory diseases such asosteoarthritis, inflammatory bowel disease, ulcerative colitis, Crohn'sdisease, idiopathic inflammatory bowel disease, irritable bowelsyndrome, spastic colon, low grade scarring (e.g., scleroderma,increased fibrosis, keloids, post-surgical scars, pulmonary fibrosis,vascular spasms, migraine, reperfusion injury and post myocardialinfarction), and sicca complex or syndrome, diseases of the centralnervous system such as stroke, pulmonary diseases such as bronchitisobliterous and primary and primary pulmonary hypertension, delayed orcell-mediated, Type IV hypersensitivity and solid and hematologicmalignancies such as leukemias and lyphomas.

In another embodiment, this invention provides a method of inhibiting anactivity of a JAK kinase, comprising contacting the JAK kinase with anamount of a compound effective to inhibit an activity of the JAK kinase,wherein the compound is selected from the compounds of this invention.In certain embodiments of the methods described herein, the method iscarried out in vivo.

In another embodiment, this invention provides a method of inhibiting anactivity of a JAK kinase, comprising contacting in vitro a JAK3 kinasewith an amount of a compound effective to inhibit an activity of the JAKkinase, wherein the compound is selected from the compounds of thisinvention.

In a specific embodiment, the compounds can be used to treat and/orprevent rejection in organ and/or tissue transplant recipients (i.e.,treat and/or prevent allorgraft rejection). Allografts can be rejectedthrough either a cell-mediated or humoral immune reaction of therecipient against transplant (histocompability) antigens present on themembranes of the donor's cells. The strongest antigens are governed by acomplex of genetic loci termed human leukocyte group A (HLA) antigens.Together with the ABO blood groups antigens, they are the chieftransplantation antigens detectable in humans.

Rejection following transplantation can generally be broken into threecategories: hyperacute, occurring hours to days followingtransplantation; acute, occurring days to months followingtransplantation; and chronic, occurring months to years followingtransplantation.

Hyperacute rejection is caused mainly by the production of hostantibodies that attack the graft tissue. In a hyperacute rejectionreaction, antibodies are observed in the transplant vascular very soonafter transplantation. Shortly thereafter, vascular clotting occurs,leading to ischemia, eventual necrosis and death. The graft infarctionis unresponsive to known immunosuppressive therapies. Because HLAantigens can be identified in vitro, pre-transplant screening is used tosignificantly reduce hyperacute rejection. As a consequence of thisscreening, hyperacute rejection is relatively uncommon today.

Acute rejection is thought to be mediated by the accumulation of antigenspecific cells in the graft tissue. The T-cell-mediated immune reactionagainst these antigens (i.e., HVGR or GVHR) is the principle mechanismof acute rejection. Accumulation of these cells leads to damage of thegraft tissue. It is believed that both CD4+ helper T-cells and CD8+cytotoxic T-cells are involved in the process and that the antigen ispresented by donor and host dendritic cells. The CD4+ helper T-cellshelp recruit other effector cells, such as macrophapges and eosinophils,to the graft. Accessing T-cell activation signal transduction cascades(for example, CD28, CD40L, and CD2 cascades) are also involved.

The cell-mediated acute rejection can be reversed in many cases byintensifying immunotherapy. After successful reversal, severely damagedelements of the graft heal by fibrosis and the remainder of the graftappears normal. After resolution of acute rejection, dosages ofimmunosuppressive drugs can be reduced to very low levels.

Chronic rejection, which is a particular problem in renal transplants,often progresses insidiously despite increased immunosuppressivetherapy. It is thought to be due, in large part, to cell-mediated TypeIV hypersensitivity. The pathologic profile differs from that of acuterejection. The arterial endothelium is primarily involved with extensiveproliferation that may gradually occlude the vessel lumen, leading toischemia, fibrosis, a thickened intima, and atherosclerotic changes.Chronic rejection is mainly due to a progressive obliteration of graftvasculature and resembles a slow, vasculitic process.

In Type IV hypersensitivity, CD8 cytotoxic T-cells and CD4 helper Tcells recognize either intracellular or extracellular synthesizedantigen when it is complexed, respectively, with either Class I or ClassII MHC molecules. Macrophages function as antigen-presenting cells andrelease IL-1, which promotes proliferation of helper T-cells. HelperT-cells release interferon gamma and IL-2, which together regulatedelayed hyperactivity reactions mediated by macrophage activation andimmunity mediated by T cells. In the case of organ transplant, thecytotoxic T-cells destroy the graft cells on contact.

Since JAK kinases play a critical role in the activation of T-cells, thecompounds described herein can be used to treat and/or prevent manyaspects of transplant rejection, and are particularly useful in thetreatment and/or prevention of rejection reactions that are mediated, atleast in part, by T-cells, such as HVGR or GVHR. The compounds can alsobe used to treat and/or prevent chronic rejection in transplantrecipients and, in particular, in renal transplant recipients. Thecompound can also be administered to a tissue or an organ prior totransplanting the tissue or organ in the transplant recipient.

In another embodiment, this invention provides a method of treating aT-cell mediated autoimmune disease, comprising administering to apatient suffering from such an autoimmune disease an amount of acompound effective to treat the autoimmune disease wherein the compoundis selected from the compounds of the invention. In certain embodimentsof the methods the autoimmune disease is multiple sclerosis (MS),psoraisis, or Sjogran's syndrome. Such autoimmune disease include, butare not limited to, those autoimmune diseases that are frequentlydesignated as single organ or single cell-type autoimmune disorders andthose autoimmune disease that are frequently designated as involvingsystemic autoimmune disorder. Non-limiting examples of diseasesfrequently designated as single organ or single cell-type autoimmunedisorders include: Hashimoto's thyroiditis, autoimmune hemolytic anemia,autoimmune atrophic gastritis of pernicious anemia, autoimmuneencephalomyelitis, autoimmune orchitis, Goodpasture's disease,autoimmune thrombocytopenia, sympathetic ophthalmia, myasthenia gravis,Graves' disease, primary biliary cirrhosis, chronic aggressivehepatitis, ulcerative colitis and membranous glomerulopathy.Non-limiting examples of diseases often designated as involving systemicautoimmune disorder include: systemic lupus erythematosis, rheumatoidarthritis, Sjogren's syndrome, Reiter's syndrome,polymyositis-dermatomyositis, systemic sclerosis, polyarteritis nodosa,multiple sclerosis and bullous pemphigoid. Additional autoimmunediseases, which can be .beta.-cell (humoral) based or T-cell based,include Cogan's syndrome, ankylosing spondylitis, Wegener'sgranulomatosis, autoimmune alopecia, Type I or juvenile onset diabetes,and thyroiditis.

The types of autoimmune diseases that may be treated or prevented withsuch prodrugs generally include those disorders involving tissue injurythat occurs as a result of a humoral and/or cell-mediated response toimmunogens or antigens of endogenous and/or exogenous origin. Suchdiseases are frequently referred to as diseases involving thenonanaphylactic (i.e., Type II, Type III and/or Type IV)hypersensitivity reactions.

Type I hypersensitivity reactions generally result from the release ofpharmacologically active substances, such as histamine, from mast and/orbasophil cells following contact with a specific exogenous antigen. Asmentioned above, such Type I reactions play a role in numerous diseases,including allergic asthma, allergic rhinitis, etc.

Type II hypersensitivity reactions (also referred to as cytotoxic,cytolytic complement-dependent or cell-stimulating hypersensitivityreactions) result when immunoglobulins react with antigenic componentsof cells or tissue, or with an antigen or hapten that has becomeintimately coupled to cells or tissue. Diseases that are commonlyassociated with Type II hypersensitivity reactions include, but are notlimited, to autoimmune hemolytic anemia, erythroblastosis fetalis andGoodpasture's disease.

Type III hypersensitivity reactions, (also referred to as toxic complex,soluble complex, or immune complex hypersensitivity reactions) resultfrom the deposition of soluble circulating antigen-immunoglobulincomplexes in vessels or in tissues, with accompanying acute inflammatoryreactions at the site of immune complex deposition. Non-limitingexamples of prototypical Type III reaction diseases include the Arthusreaction, rheumatoid arthritis, serum sickness, systemic lupuserythematosis, certain types of glomerulonephritis, multiple sclerosisand bullous pemphingoid.

Type IV hypersensitivity reactions (frequently called cellular,cell-mediated, delayed, or tuberculin-type hypersensitivity reactions)are caused by sensitized T-lymphocytes which result from contact with aspecific antigen. Non-limiting examples of diseases cited as involvingType IV reactions are contact dermatitis and allograft rejection.

Autoimmune diseases associated with any of the above nonanaphylactichypersensitivity reactions may be treated or prevented with the prodrugsaccording to structural formulae (I) and (Ia). In particular, themethods may be used to treat or prevent those autoimmune diseasesfrequently characterized as single organ or single cell-type autoimmunedisorders including, but not limited to: Hashimoto's thyroiditis,autoimmune hemolytic anemia, autoimmune atrophic gastritis of perniciousanemia, autoimmune encephalomyelitis, autoimmune orchitis, Goodpasture'sdisease, autoimmune thrombocytopenia, sympathetic ophthalmia, myastheniagravis, Graves' disease, primary biliary cirrhosis, chronic aggressivehepatitis, ulcerative colitis and membranous glomerulopathy, as well asthose autoimmune diseases frequently characterized as involving systemicautoimmune disorder, which include but are not limited to: systemiclupus erythematosis (SLE), rheumatoid arthritis, Sjogren's syndrome,Reiter's syndrome, polymyositis-dermatomyositis, systemic sclerosis,polyarteritis nodosa, multiple sclerosis and bullous pemphigoid.

It will be appreciated by skilled artisans that many of the above-listedautoimmune diseases are associated with severe symptoms, theamelioration of which provides significant therapeutic benefit even ininstances where the underlying autoimmune disease may not beameliorated.

Therapy using the compounds described herein can be applied alone, or itcan be applied in combination with or adjunctive to other commonimmunosuppressive therapies, such as, for example, the following:mercaptopurine; corticosteroids such as prednisone; methylprednisoloneand prednisolone; alkylating agents such as cyclophosphamide;calcineurin inhibitors such as cyclosporine, sirolimus, and tacrolimus;inhibitors of inosine monophosphate dehydrogenase (IMPDH) such asmycophenolate, mycophenolate mofetil, and azathioprine; and agentsdesigned to suppress cellular immunity while leaving the recipient'shumoral immunologic response intact, including various antibodies (forexample, antilymphocyte globulin (ALG), antithymocyte globulin (ATG),monoclonal anti-T-cell antibodies (OKT3)) and irradiation. These variousagents can be used in accordance with their standard or common dosages,as specified in the prescribing information accompanying commerciallyavailable forms of the drugs (see also: the prescribing information inthe 2006 Edition of The Physician's Desk Reference), the disclosures ofwhich are incorporated herein by reference. Azathioprine is currentlyavailable from Salix Pharmaceuticals, Inc., under the brand name AZASAN;mercaptopurine is currently available from Gate Pharmaceuticals, Inc.,under the brand name PURINETHOL; prednisone and prednisolone arecurrently available from Roxane Laboratories, Inc.; Methyl prednisoloneis currently available from Pfizer; sirolimus (rapamycin) is currentlyavailable from Wyeth-Ayerst under the brand name RAPAMUNE; tacrolimus iscurrently available from Fujisawa under the brand name PROGRAF;cyclosporine is current available from Novartis under the brand dameSANDIMMUNE and from Abbott under the brand name GENGRAF; IMPDHinhibitors such as mycophenolate mofetil and mycophenolic acid arecurrently available from Roche under the brand name CELLCEPT and fromNovartis under the brand name MYFORTIC; azathioprine is currentlyavailable from Glaxo Smith Kline under the brand name IMURAN; andantibodies are currently available from Ortho Biotech under the brandname QRTHOCLONE, from Novartis under the brand name SIMULECT(basiliximab), and from Roche under the brand name ZENAPAX (daclizumab).

In another embodiment, the compounds could be administered either incombination or adjunctively with an inhibitor of a syk kinase. sykkinase is a tyrosine kinase known to play a critical role in Fcyreceptor signaling, as well as in other signaling cascades, such asthose involving B-cell receptor signaling (Turner et al., (2000),Immunology Today 21:148-154) and integrins beta (1), beta (2), and beta(3) in neutrophils (Mocsai et al., (2002), Immunity 16:547-558). Forexample, syk kinase plays a pivotal role in high affinity IgE receptorsignaling in mast cells that leads to activation and subsequent releaseof multiple chemical mediators that trigger allergic attacks. However,unlike the JAK kinases, which help regulate the pathways involved indelayed or cell-mediated Type IV hypersensitivity reactions, syk kinasehelps regulate the pathways involved in immediate IgE-mediated, Type Ihypersensitivity reactions. Certain compounds that affect the sykpathway may or may not also affect the JAK pathways.

Suitable syk inhibitory compounds are described, for example, in Ser.No. 10/355,543 filed Jan. 31, 2003 (publication no. 2004/0029902); WO03/063794; Ser. No. 10/631,029 filed Jul. 29, 2003; WO 2004/014382; Ser.No. 10/903,263 filed Jul. 30, 2004; PCT/US2004/24716 filed Jul. 30, 2004(WO005/016893); Ser. No. 10/903,870 filed Jul. 30, 2004;PCT/US2004/24920 filed Jul. 30, 2004; Ser. No. 60/630,808 filed Nov. 24,2004; Ser. No. 60/645,424 filed Jan. 19, 2005; and Ser. No. 60/654,620,filed Feb. 18, 2005, the disclosures of which are incorporated herein byreference. The described herein and syk inhibitory compounds could beused alone or in combination with one or more conventional transplantrejection treatments, as described above.

In a specific embodiment, the compounds can be used to treat or preventthese diseases in patients that are either initially non-responsive(resistant) to or that become non-responsive to treatment with a sykinhibitory compound or one of the other current treatments for theparticular disease. The compounds could also be used in combination withsyk inhibitory compounds in patients that are syk-compound resistant ornon-responsive. Suitable syk-inhibitory compounds with which thecompounds can be administered are provided infra.

In another embodiment, this invention provides a method of treating aT-cell mediated autoimmune disease, comprising administering to apatient suffering from such an autoimmune disease an amount of acompound effective to treat the autoimmune disease wherein the compoundis selected from the compounds of the invention, as described herein,and the compound is administered in combination with or adjunctively toa compound that inhibits syk kinase with an IC₅₀ in the range of atleast 10 μM.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient,comprising administering to the transplant recipient an amount of acompound effective to treat or prevent the rejection wherein thecompound is selected from the compounds of the invention, as describedherein. In a further embodiment, the compound is administered to atissue or an organ prior to transplanting the tissue or organ in thetransplant recipient.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is acute rejection, comprising administering to thetransplant recipient an amount of a compound effective to treat orprevent the rejection, wherein the compound is selected from thecompounds of the invention.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is chronic rejection, comprising administering tothe transplant recipient an amount of a compound effective to treat orprevent the rejection, wherein the compound is selected from thecompounds of the invention.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is mediated by HVGR or GVHR, comprisingadministering to the transplant recipient an amount of a compoundeffective to treat or prevent the rejection, wherein the compound isselected from the compounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejection,wherein the compound is selected from the compounds of this invention,as described herein.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejectionwherein the compound is selected from the compounds of the invention, asdescribed herein, in which the compound is administered in combinationwith or adjunctively to another immunosuppressant.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejection,wherein the compound is selected from the compounds of the invention, asdescribed herein, in which the compound is administered in combinationwith or adjunctively to another immunosuppressant, in which theimmunosuppressant is selected from cyclosporine, tacrolimus, sirolimus,an inhibitor of IMPDH, mycophenolate, mycophanolate mofetil, ananti-T-Cell antibody, and OKT3.

The compounds described herein are cytokine moderators of IL-4signaling. As a consequence, the compounds could slow the response ofType I hypersensitivity reactions. Thus, in a specific embodiment, thecompounds could be used to treat such reactions and, therefore, thediseases associated with, mediated by, or caused by suchhypersensitivity reactions (for example, allergies), prophylactically.For example, an allergy sufferer could take one or more of the JAKselective compounds described herein prior to expected exposure toallergens to delay the onset or progress of, or eliminate altogether, anallergic response.

When used to treat or prevent such diseases, the compounds can beadministered singly, as mixtures of one or more compounds, or in mixtureor combination with other agents useful for treating such diseasesand/or the symptoms associated with such diseases. The compounds mayalso be administered in mixture or in combination with agents useful totreat other disorders or maladies, such as steroids, membranestabilizers, 5-lipoxygenase (5LO) inhibitors, leukotriene synthesis andreceptor inhibitors, inhibitors of IgE isotype switching or IgEsynthesis, IgG isotype switching or IgG synthesis, beta.-agonists,tryptase inhibitors, aspirin, cyclooxygenase (COX) inhibitors,methotrexate, anti-TNF drugs, anti CD20 antibody, PD4 inhibitors, p38inhibitors, PDE4 inhibitors, and antihistamines, to name a few. Thecompounds can be administered per se in the form of prodrugs or aspharmaceutical compositions, comprising an active compound or prodrug.

In another embodiment, this invention provides a method of treating orpreventing a Type IV hypersensitivity reaction, comprising administeringto a subject an amount of a compound effective to treat or prevent thehypersensitivity reaction, wherein the compound is selected from thecompounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a Type IV hypersensitivity reaction, which is practicalprophylactically, comprising administering to a subject an amount of acompound effective to treat or prevent the hypersensitivity reaction,wherein the compound is selected from the compounds of this invention,as described herein, and is administered prior to exposure to anallergen.

In another embodiment, this invention provides a method of inhibiting asignal transduction cascade in which JAK3 kinase plays a role,comprising contacting a cell expressing a receptor involved in such asignaling cascade with a compound wherein the compound is selected fromthe compounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, comprising administering to asubject an amount of compound effective to treat or prevent the JAKkinase-mediated disease, wherein the compound is selected from thecompounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, in which the JAK-mediateddisease is HVGR or GVHR, comprising administering to a subject an amountof compound effective to treat or prevent the JAK kinase-mediateddisease, wherein the compound is selected from the compounds of theinvention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, in which the JAK-mediateddisease is acute allograft rejection, comprising administering to asubject an amount of compound effective to treat or prevent the JAKkinase-mediated disease, wherein the compound is selected from thecompounds of the invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a syk kinase-mediated disease, in which the JAK-mediateddisease is chronic allograft rejection, comprising administering to asubject an amount of compound effective to treat or prevent the JAKkinase-mediated disease, wherein the compound is selected from thecompounds of the invention, as described herein.

Active compounds of the invention typically inhibit the syk pathway. Theactivity of a specified compound as an inhibitor of a syk kinase can beassessed in vitro or in vivo. In some embodiments, the activity of aspecified compound can be tested in a cellular assay.

“Cell proliferative disorder” refers to a disorder characterized byabnormal proliferation of cells. A proliferative disorder does not implyany limitation with respect to the rate of cell growth, but merelyindicates loss of normal controls that affect growth and cell division.Thus, in some embodiments, cells of a proliferative disorder can havethe same cell division rates as normal cells but do not respond tosignals that limit such growth. Within the ambit of “cell proliferativedisorder” is neoplasm or tumor, which is an abnormal growth of tissue.Cancer refers to any of various malignant neoplasms characterized by theproliferation of cells that have the capability to invade surroundingtissue and/or metastasize to new colonization sites.

Generally, cell proliferative disorders treatable with the compoundsdisclosed herein relate to any disorder characterized by aberrant cellproliferation. These include various tumors and cancers, benign ormalignant, metastatic or non-metastatic. Specific properties of cancers,such as tissue invasiveness or metastasis, can be targeted using themethods described herein. Cell proliferative disorders include a varietyof cancers, including, among others, ovarian cancer, renal cancer,gastrointestinal cancer, kidney cancer, bladder cancer, pancreaticcancer, lung squamous carcinoma, and adenocarcinoma.

In some embodiments, the cell proliferative disorder treated is ahematopoietic neoplasm, which is aberrant growth of cells of thehematopoietic system. Hematopoietic malignancies can have its origins inpluripotent stem cells, multipotent progenitor cells, oligopotentcommitted progenitor cells, precursor cells, and terminallydifferentiated cells involved in hematopoiesis. Some hematologicalmalignancies are believed to arise from hematopoietic stem cells, whichhave the ability for self renewal. For instance, cells capable ofdeveloping specific subtypes of acute myeloid leukemia (AML) (Cynthia K.Hahn, Kenneth N. Ross, Rose M. Kakoza, Steven Karr, Jinyan Du, Shao-EOng, Todd R. Golub, Kimberly Stegmaier, Syk is a new target for AMLdifferentiation, Blood, 2007, 110, Abstract 209) upon transplantationdisplay the cell surface markers of hematopoietic stem cells,implicating hematopoietic stem cells as the source of leukemic cells.Blast cells that do not have a cell marker characteristic ofhematopoietic stem cells appear to be incapable of establishing tumorsupon transplantation (Blaire et al., 1997, Blood 89:3104-3112). The stemcell origin of certain hematological malignancies also finds support inthe observation that specific chromosomal abnormalities associated withparticular types of leukemia can be found in normal cells ofhematopoietic lineage as well as leukemic blast cells. For instance, thereciprocal translocation t(9q34;22q11) associated with approximately 95%of chronic myelogenous leukemia appears to be present in cells of themyeloid, erythroid, and lymphoid lineage, suggesting that thechromosomal aberration originates in hematopoietic stem cells. Asubgroup of cells in certain types of CML displays the cell markerphenotype of hematopoietic stem cells.

Although hematopoietic neoplasms often originate from stem cells,committed progenitor cells or more terminally differentiated cells of adevelopmental lineage can also be the source of some leukemias. Forexample, forced expression of the fusion protein Bcr/Abl (associatedwith chronic myelogenous leukemia) in common myeloid progenitor orgranulocyte/macrophage progenitor cells produces a leukemic-likecondition. Moreover, some chromosomal aberrations associated withsubtypes of leukemia are not found in the cell population with a markerphenotype of hematopoietic stem cells, but are found in a cellpopulation displaying markers of a more differentiated state of thehematopoietic pathway (Turhan et al., 1995, Blood 85:2154-2161). Thus,while committed progenitor cells and other differentiated cells may haveonly a limited potential for cell division, leukemic cells may haveacquired the ability to grow unregulated, in some instances mimickingthe self-renewal characteristics of hematopoietic stem cells (Passegueet al., Proc. Natl. Acad. Sci. USA, 2003, 100:11842-9).

In some embodiments, the hematopoietic neoplasm treated is a lymphoidneoplasm, where the abnormal cells are derived from and/or display thecharacteristic phenotype of cells of the lymphoid lineage. Lymphoidneoplasms can be subdivided into B-cell neoplasms, T and NK-cellneoplasms, and Hodgkin's lymphoma. B-cell neoplasms can be furthersubdivided into precursor B-cell neoplasm and mature/peripheral B-cellneoplasm. Exemplary B-cell neoplasms are precursor B-lymphoblasticleukemia/lymphoma (precursor B-cell acute lymphoblastic leukemia) whileexemplary mature/peripheral B-cell neoplasms are B-cell chroniclymphocytic leukemia/small lymphocytic lymphoma, B-cell prolymphocyticleukemia, lymphoplasmacytic lymphoma, splenic marginal zone B-celllymphoma, hairy cell leukemia, plasma cell myeloma/plasmacytoma,extranodal marginal zone B-cell lymphoma of MALT type, nodal marginalzone B-cell lymphoma, follicular lymphoma, mantle-cell lymphoma, diffuselarge B-cell lymphoma, mediastinal large B-cell lymphoma, primaryeffusion lymphoma, and Burkitt's lymphoma/Burkitt cell leukemia. T-celland Nk-cell neoplasms are further subdivided into precursor T-cellneoplasm and mature (peripheral) T-cell neoplasms. Exemplary precursorT-cell neoplasm is precursor T-lymphoblastic lymphoma/leukemia(precursor T-cell acute lymphoblastic leukemia) while exemplary mature(peripheral) T-cell neoplasms are T-cell prolymphocytic leukemia T-cellgranular lymphocytic leukemia, aggressive NK-cell leukemia, adult T-celllymphoma/leukemia (HTLV-1), extranodal NK/T-cell lymphoma, nasal type,enteropathy-type T-cell lymphoma, hepatosplenic gamma-delta T-celllymphoma, subcutaneous panniculitis-like T-cell lymphoma, Mycosisfungoides/Sezary syndrome, Anaplastic large-cell lymphoma, T/null cell,primary cutaneous type, Peripheral T-cell lymphoma, not otherwisecharacterized, Angioimmunoblastic T-cell lymphoma, Anaplastic large-celllymphoma, T/null cell, primary systemic type. The third member oflymphoid neoplasms is Hodgkin's lymphoma, also referred to as Hodgkin'sdisease. Exemplary diagnosis of this class that can be treated with thecompounds include, among others, nodular lymphocyte-predominantHodgkin's lymphoma, and various classical forms of Hodgkin's disease,exemplary members of which are Nodular sclerosis Hodgkin's lymphoma(grades 1 and 2), Lymphocyte-rich classical Hodgkin's lymphoma, Mixedcellularity Hodgkin's lymphoma, and Lymphocyte depletion Hodgkin'slymphoma. In various embodiments, any of the lymphoid neoplasms that areassociated with aberrant JAK activity can be treated with the sykinhibitory compounds.

In some embodiments, the hematopoietic neoplasm treated is a myeloidneoplasm. This group comprises a large class of cell proliferativedisorders involving or displaying the characteristic phenotype of thecells of the myeloid lineage. Myeloid neoplasms can be subdivided intomyeloproliferative diseases, myelodysplastic/myeloproliferativediseases, myelodysplastic syndromes, and acute myeloid leukemias.Exemplary myeloproliferative diseases are chronic myelogenous leukemia(e.g., Philadelphia chromosome positive (t(9;22)(qq34;q11)), chronicneutrophilic leukemia, chronic eosinophilic leukemia/hypereosinophilicsyndrome, chronic idiopathic myelofibrosis, polycythemia vera, andessential thrombocythemia. Exemplary myelodysplastic/myeloproliferativediseases are chronic myelomonocytic leukemia, atypical chronicmyelogenous leukemia, and juvenile myelomonocytic leukemia. Exemplarymyelodysplastic syndromes are refractory anemia, with ringedsideroblasts and without ringed sideroblasts, refractory cytopenia(myelodysplastic syndrome) with multilineage dysplasia, refractoryanemia (myelodysplastic syndrome) with excess blasts, 5q-syndrome, andmyelodysplastic syndrome. In various embodiments, any of the myeloidneoplasms that are associated with aberrant syk activity can be treatedwith the syk inhibitory compounds.

In some embodiments, the compounds can be used to treat Acute myeloidleukemias (AML), which represent a large class of myeloid neoplasmshaving its own subdivision of disorders. These subdivisions include,among others, AMLs with recurrent cytogenetic translocations, AML withmultilineage dysplasia, and other AML not otherwise categorized.Exemplary AMLs with recurrent cytogenetic translocations include, amongothers, AML with t(8;21)(q22;q22), AML1(CBF-alpha)/ETO, Acutepromyelocytic leukemia (AML with t(15;17)(q22;q11-12) and variants,PML/RAR-alpha), AML with abnormal bone marrow eosinophils(inv(16)(p13q22) or t(16;16)(p13;q11), CBFb/MYH11X), and AML with 11q23(MLL) abnormalities. Exemplary AML with multilineage dysplasia are thosethat are associated with or without prior myelodysplastic syndrome.Other acute myeloid leukemias not classified within any definable groupinclude, AML minimally differentiated, AML without maturation, AML withmaturation, Acute myelomonocytic leukemia, Acute monocytic leukemia,Acute erythroid leukemia, Acute megakaryocytic leukemia, Acutebasophilic leukemia, and Acute panmyelosis with myelofibrosis.

“Treating” within the context of the invention means an alleviation ofsymptoms associated with a disorder or disease, or halt of furtherprogression or worsening of those symptoms, or prevention or prophylaxisof the disease or disorder.

The term “mammal” includes organisms which express syk. Examples ofmammals include mice, rats, cows, sheep, pigs, goats, horses, bears,monkeys, dogs, cats and, preferably, humans. Transgenic organisms whichexpress syk are also included in this definition.

The inventive methods comprise administering an effective amount of acompound or composition described herein to a mammal or non-humananimal. As used herein, “effective amount” of a compound or compositionof the invention includes those amounts that antagonize or inhibit syk.An amount which antagonizes or inhibits syk is detectable, for example,by any assay capable of determining syk activity, including the onedescribed below as an illustrative testing method. Effective amounts mayalso include those amounts which alleviate symptoms of a syk associateddisorder treatable by inhibiting syk. Accordingly, “antagonists of syk”include compounds which interact with the syk, and modulate, e.g.,inhibit or decrease, the ability of a second compound, e.g., another sykligand, to interact with the syk. The syk binding compounds arepreferably antagonists of syk. The language “syk binding compound”(e.g., exhibits binding affinity to the receptor) includes thosecompounds which interact with syk resulting in modulation of theactivity of syk. Syk binding compounds may be identified using an invitro (e.g., cell and non-cell based) or in vivo method. A descriptionof in vitro methods are provided below.

The amount of compound present in the methods and compositions describedherein should be sufficient to cause a detectable decrease in theseverity of the disorder, as measured by any of the assays described inthe examples. The amount of syk modulator needed will depend on theeffectiveness of the modulator for the given cell type and the length oftime required to treat the disorder. In certain embodiments, thecompositions of this invention may further comprise another therapeuticagent. When a second agent is used, the second agent may be administeredeither as a separate dosage form or as part of a single dosage form withthe compounds or compositions of this invention. While one or more ofthe inventive compounds can be used in an application of monotherapy totreat a disorder, disease or symptom, they also may be used incombination therapy, in which the use of an inventive compound orcomposition (therapeutic agent) is combined with the use of one or moreother therapeutic agents for treating the same and/or other types ofdisorders, symptoms and diseases. Combination therapy includesadministration of the two or more therapeutic agents concurrently orsequentially. The agents may be administered in any order.Alternatively, the multiple therapeutic agents can be combined into asingle composition that can be administered to the patient. Forinstance, a single pharmaceutical composition could comprise thecompound or pharmaceutically acceptable salt, ester or prodrug thereofaccording to the formula I, another therapeutic agent (e.g.,methotrexate) or a tautomer thereof or a pharmaceutically acceptablesalt, ester or prodrug thereof, and a pharmaceutically acceptableexcipient or carrier.

The invention comprises a compound having the formula I, a method formaking an inventive compound, a method for making a pharmaceuticalcomposition from at least one inventive compound and at least onepharmaceutically acceptable carrier or excipient, and a method of usingone or more inventive compounds to treat a variety of disorders,symptoms and diseases (e.g., inflammatory, autoimmune, neurological,neurodegenerative, oncology and cardiovascular), such as RA,osteoarthritis, irritable bowel disease IBD, asthma, chronic obstructivepulmonary disease COPD and MS. The inventive compounds and theirpharmaceutically acceptable salts and/or neutral compositions may beformulated together with a pharmaceutically acceptable excipient orcarrier and the resulting composition may be administered in vivo tomammals, such as men, women and animals, to treat a variety ofdisorders, symptoms and diseases. Furthermore, the inventive compoundscan be used to prepare a medicament that is useful for treating avariety of disorders, symptoms and diseases.

All of the compounds of the present invention are either potentinhibitors of syk kinases, exhibiting IC₅₀s in the respective assay inthe range of less than 5 μM, with most being in the nanomolar, andseveral in the sub-nanomolar, range. In some embodiments, the compoundsof the present invention may be “dual” syk/JAK inhibitors in that theyinhibit both syk and JAK kinase to some degree. In other embodiments,the compounds of the present invention may selectively inhibit sykkinase, but not appreciably inhibit one or more JAK kinases. In otherembodiments, the compounds of the present invention may selectivelyinhibit JAK kinase, but not appreciably inhibit one or more syk kinases.

f. Kits

Still another aspect of this invention is to provide a kit comprisingseparate containers in a single package, wherein the inventivepharmaceutical compounds, compositions and/or salts thereof are used incombination with pharmaceutically acceptable carriers to treat states,disorders, symptoms and diseases where syk plays a role.

I. EXAMPLES

The following examples are offered to illustrate, but not to limit, theclaimed invention.

The starting materials and reagents used in preparing these compoundsgenerally are either available from commercial suppliers, such asAldrich Chemical Co., or are prepared by methods known to those skilledin the art following procedures set forth in references such as Fieserand Fieser's Reagents for Organic Synthesis; Wiley & Sons: New York,1967-2004, Volumes 1-22; Rodd's Chemistry of Carbon Compounds, ElsevierScience Publishers, 1989, Volumes 1-5 and Supplementals; and OrganicReactions, Wiley & Sons: New York, 2005, Volumes 1-65.

The starting materials and the intermediates of the synthetic reactionschemes can be isolated and purified if desired using conventionaltechniques, including but not limited to, filtration, distillation,crystallization, chromatography, and the like. Such materials can becharacterized using conventional means, including physical constants andspectral data.

Unless specified to the contrary, the reactions described hereinpreferably are conducted under an inert atmosphere at atmosphericpressure at a reaction temperature range from about −78° C. to about150° C., more preferably from about 0° C. to about 125° C., and mostpreferably and conveniently at about room (or ambient) temperature,e.g., about 20° C. to about 75° C.

Referring to the examples that follow, compounds of the presentinvention were synthesized using the methods described herein, or othermethods, which are well known in the art.

The compounds and/or intermediates may be characterized by highperformance liquid chromatography (HPLC) using a Waters Alliancechromatography system with a 2695 Separation Module (Milford, Mass.).The analytical columns may be C-18 SpeedROD RP-18E Columns from MerckKGaA (Darmstadt, Germany). Alternately, characterization may beperformed using a Waters Unity (HPLC) system with Waters Acquity HPLCBEH C-18 2.1 mm×15 mm columns. A gradient elution may be used, typicallystarting with 5% acetonitrile/95% water and progressing to 95%acetonitrile over a period of 5 minutes for the Alliance system and 1minute for the Acquity system. All solvents may contain 0.1%trifluoroacetic acid (TFA). Compounds may be detected by ultravioletlight (UV) absorption at either 220 or 254 nm. HPLC solvents may be fromEMD Chemicals, Inc. (Gibbstown, N.J.). In some instances, purity may beassessed by thin layer chromatography (TLC) using glass backed silicagel plates, such as, for example, EMD Silica Gel 60 2.5 cm×7.5 cmplates. TLC results may be readily detected visually under ultravioletlight, or by employing well known iodine vapor and other variousstaining techniques.

Mass spectrometric analysis may be performed on one of two Agilent 1100series LCMS instruments and the Acquity system with acetonitrile/wateras the mobile phase. One system may use TFA as the modifier and measurein positive ion mode [reported as MH+, (M+1) or (M+H)+] and the othermay use either formic acid or ammonium acetate and measure in bothpositive [reported as MH⁺, (M+1) or (M+H)⁺] and negative [reported asM−, (M−1) or (M−H)⁻] ion modes.

Nuclear magnetic resonance (NMR) analysis may be performed on some ofthe compounds with a Varian 400 MHz NMR (Palo Alto, Calif.). Thespectral reference may be either TMS or the known chemical shift of thesolvent.

The purity of some of the invention compounds may be assessed byelemental analysis (Robertson Microlit, Madison N.J.).

Melting points may be determined on a Laboratory Devices MeI-Tempapparatus (Holliston, Mass.).

Preparative separations may be carried out as needed, using either anSql 6× or an Sg100c chromatography system and prepackaged silica gelcolumns all purchased from Teledyne Isco, (Lincoln, Nebr.). Alternately,compounds and intermediates may be purified by flash columnchromatography using silica gel (230-400 mesh) packing material, or byHPLC using a C-18 reversed phase column. Typical solvents employed forthe Isco systems and flash column chromatography may be dichloromethane,methanol, ethyl acetate, hexane, acetone, aqueous hydroxyamine andtriethyl amine. Typical solvents employed for the reverse phase HPLC maybe varying concentrations of acetonitrile and water with 0.1%trifluoroacetic acid.

General Methods

The following synthetic reaction schemes are merely illustrative of somemethods by which the compounds of the present invention can besynthesized, and various modifications to these synthetic reactionschemes can be made and will be suggested to one skilled in the arthaving referred to the disclosure contained in this application.

Example 14-(4-(4-acetylpiperazin-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

Step 1: To a stirring solution of carboxylic acid 72.1 (85 g, 540 mmol)in thionyl chloride (425 mL) was added pyridine (8.5 mL, 0.11 mmol),slowly. The reaction was stirred at 75° C. overnight at which time itwas concentrated and dried under vacuum to a light yellow powder. Thisyellow solid was slowly diluted with 750 mL of ethanol and refluxedovernight. The next day the reaction was determined to be complete byHPLC and then cooled in an ice bath and the solid filtered and washedwith diethyl ether affording ethyl ester 72.2 an off-white powder (91 g,87% for two steps). MS found for C₇H₈N₂O₄ as (M+H)⁺ 185.0.

Step 2: Ester 72.2 (22 g, 120 mmol) was dissolved in phosphorousoxychloride (60 mL, 600 mmol) and the mixture treated withN,N-diethylaniline (27 mL, 167 mmol) and the mixture heated to 105° C.until the reaction was determined to be complete by HPLC. It was thencooled to RT and slowly added to 1 L of crushed ice resulting in theformation of a beige precipitate which was collected by filtration anddried under vacuum affording dichloride 72.3a light yellow powder (22.5g, 85%). ¹H NMR (DMSO-d₆, 400 MHz): δ 9.13 (s, 1H), 4.37 (q, 2H), 1.32(t, 3H).

Step 3: Dichloropyrimidine 72.3 (500 mg, 2.3 mmol) was dissolved in NMP(10 mL) and stirred in ice bath. To it was added a soluntion of aniline72.4 (540 mg, 2.5 mmol) and ethyldiisopropylamine (DIEA, 0.82 mL, 4.6mmol) in 10 mL NMP dropwise using an additional funnel. The mixture wasstirred for 1 hour, diluted with ethyl acetate, washed with brine,concentrated and subjected to flash column to isolate compound 72.5 awhite solid (905 mg, 97%). MS found for C₁₉H₂₂ClN₅O₃ as (M+H)⁺ 404.1.

Step 4: Ethyl ester 72.5 (5 mg, 2.2 mmol) was dissolved in 100 mL THF.To it were added lithium hydroxide hydrate (190 mg, 4.4 mmol) and 10 mLwater. The mixture was stirred for 1 hour and to it was carefully added1N HCl solution till pH reaching 3. The mixture was concentrated invacuo to remove THF. White solid crashed out and was isolated using aBüd chner funnel. It was washed with water and dried in vacuum oven togive compound 72.6 (900 mg, 99%) as a white solid. MS found forC₁₇H₁₈ClN₅O₃ as (M+H)⁺ 376.1.

Step 5: Carboxylic acid 72.6 (900 mg, 2.2 mmol) was dissolved in 30 mLNMP. To it were added EDC hydrochloride (690 mg, 3.6 mmol) and HOBthydrate (490 mg, 3.6 mmol). The mixture was stirred at RT for 90minutes. To it was then added ammonia (commercial 0.5N solution indioxane, 24 mL, 12 mmol). The mixture was stirred for overnight. It wasthen concentrated in vacuo and taken into water and chloroform. Thechloroform phase was separated and washed with brine four times. Thechloroform phase was then dried over MgSO₄ and concentrated in vacuo toafford compound 72.7 as a light yellow solid (720 mg, 63%). MS found forC₂₃H₂₃N₉O₃ as (M+H)⁺ 474.2.

Step 6: Benzotriazolyl ether 72.7 (100 mg, 0.21 mmol) was dissolved in 3mL DMSO. To it was added cis-1,2-diaminocyclohexane (124 μL, 1.05 mmol).The mixture was stirred for 1 hour at 120° C. bath in a sealed flask.This mixture was then subjected to preparative HPLC to isolate theracemic title compound 72. MS found for C₂₃H₃₂N₈O₂ as (M+H)⁺ 453.2. UVλ=240, 297 nm.

Example 22-((1R,2S)-2-aminocyclohexylamino)-4-(4-(1,1-dioxo)thiomorpholinophenylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 1 with 4-(1,1-dioxo)thiomorpholinoaniline toreplace aniline 72.4. MS found for C₂₁H₂₉N₇O₃S as (M+H)⁺ 460.2. UVλ=236, 312 nm.

Example 34-(4-(1H-pyrazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

Step 1: Dichloropyrimidine 72.3 (1.12 g, 5.05 mmol) was dissolved in 60mL DMF and stirred at RT. To it were added 4-(1H-pyrazol-1-yl)aniline74.1 (0.96 g, 6.1 mmol) in one portion and ethyldiisopropylamine (DIEA,1.58 mL, 9.1 mmol) dropwise using a syringe. The mixture was stirred for1 hour at RT. To it was then added sodium thiomethxoide (885 mg, 12.6mmol). The mixture was stirred at RT for overnight, diluted with ethylacetate, washed with brine three times, dried over MgSO₄, concentratedin vacuo and subjected to flash column to afford compound 50.2 was awhite solid. MS found for C₁₇H₁₇N₅O₂S as (M+H)⁺ 356.1.

Step 2: Ethyl ester 74.2 from Step 1 was dissolved in 200 mL THF. To itwere added lithium hydroxide hydrate (424 mg, 10.1 mmol) and 20 mLwater. The mixture was stirred for 2 days at RT. It was concentrated invacuo to remove THF and carefully treated with 1N HCl till pH reaching3. A white solid crashed out from the solution. It was isolated using aBüchner funnel, washed with cold water, dried in vacuum oven to givecompound 74.3 (1.12 g, 68% for 2 step 2). MS found for C₁₅H₁₃N₅O₂S as(M+H)⁺ 328.1.

Step 3: Carboxylic acid 74.3 (1.12 g, 3.4 mmol) was dissolved in 60 mLDMF. To it were added EDC hydrochloride (0.98 g, 5.1 mmol) and HOBthydrate (0.69 g, 5.1 mmol). The mixture was stirred at RT for 2 hours.To it was then added ammonia (commercial 0.5N solution in dioxane, 34mL, 17 mmol). The mixture was stirred for 2 hours. It was thenconcentrated in vacuo to remove dioxane. To it was added 300 mL water. Alight yellow solid crashed out. It was isolated using a Büchner funnel,washed with cold water, dried in vacuum oven to give compound 74.4 (1.12g, 99%). MS found for C₁₅H₁₄N₆OS as (M+H)⁺ 327.1.

Step 4: Compound 74.4 (50 mg, 0.15 mmol) was dissolved in 4 mL NMP. Toit was added MCPBA (65% pure, 45 mg, 0.17 mmol). It was stirred at RTfor 30 minutes. To it then were added cis-1,2-diaminocyclohexane (53 μL,0.45 mmol) and DIEA (78 μL, 0.45 mmol). The mixture was stirred for 50minutes at 90° C. bath. This mixture was then subjected to preparativeHPLC to isolate the racemic title compound 74. MS found for C₂₀H₂₄N₈O as(M+H)⁺ 393.2. UV λ=239, 304 nm. NMR (CD₃OD): δ 8.41 (s, 1H), 8.12 (m,1H), 7.70 (d, J=8.4 Hz, 2H), 7.63 (d, J=7.2 Hz, 2H), 7.62 (s, 1H), 6.43(dd, J=2.4, 2.0 Hz, 1H), 4.27 (m, 1H), 3.60 (m, 1H), 1.83-1.45 (m, 8H)ppm.

Example 44-(4-(1,2,3-thiadiazol-4-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 3 with 4-(1,2,3-thiadiazol-4-yl)aniline toreplace aniline 74.1. MS found for C₁₉H₂₂N₈OS as (M+H)⁺ 411.2. UV λ=231,311 nm. NMR (CD₃OD): δ 9.24 (s, 1H), 8.56 (s, 1H), 8.15 (d, J=5.6 Hz,2H), 7.84 (m, 2H), 4.43 (m, 1H), 3.76 (m, 1H), 1.91-1.60 (m, 8H) ppm.

Example 52-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridin-4-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Dichloropyrimidine 72.3 (3.02 g, 13.6 mmol) was dissolved in 100mL DMF and stirred at RT. To it were added 4-iodoaniline 77.1 (3.59 g,16.3 mmol) in one portion and ethyldiisopropylamine (DIEA, 4.25 mL, 24.4mmol) dropwise using a syringe. The mixture was stirred for 2.5 hours atRT. To it was then added sodium thiomethxoide (2.38 g, 34 mmol). Themixture was stirred at RT for 1.5 hr, diluted with ethyl acetate, washedwith brine three times, dried over MgSO₄, treated with activated carbon,concentrated in vacuo to afford crude compound 77.2 was a light brownsolid (4.50 g, 79%). MS found for C₁₄H₁₄IN₃O₂S as (M+H)⁺ 416.1.

Step 2: Iodobenzene 77.2 (620 mg, 1.5 mmol) was dissolved in 20 mLdioxane and 10 mL water. To it were added boronic acid 77.3 (406 mg, 3.3mmol), Na₂CO₃ (480 mg, 4.5 mmol) and Pd(PPh₃)₂Cl₂ (211 mg, 0.3 mmol).The mixture was degassed using argon stream for 3 minutes and stirred in85° C. bath under argon for 1 hour. The mixture was concentrated anddiluted with chloroform. It was washed with brine, dried over MgSO₄,concentrated and subjected to flash column to isolate compound 77.4 (300mg, 55%). MS found for C₁₉H₁₈N₄O₂S as (M+H)⁺ 367.1.

Step 3: Ethyl ester 77.4 (300 mg, 0.82 mmol) was dissolved in 30 mL THF.To it were added lithium hydroxide hydrate (104 mg, 2.46 mmol) and 10 mLwater. The mixture was stirred for 2 hours at RT. It was concentrated invacuo to remove THF and carefully treated with 1N HCl till pH reaching3. A light yellow solid crashed out from the solution. It was isolatedusing a Büchner funnel, washed with cold water, dried in vacuum oven togive compound 77.5 (250 mg, 90%). MS found for C₁₇H₁₄N₄O₂S as (M+H)⁺339.1.

Step 4: Carboxylic acid 77.5 (250 mg, 0.74 mmol) was dissolved in 10 mLDMF. To it were added EDC hydrochloride (213 mg, 1.1 mmol) and HOBthydrate (150 mg, 1.1 mmol). The mixture was stirred at RT for 90minutes. To it was then added ammonia (commercial 0.5N solution indioxane, 7.4 mL, 3.7 mmol). The mixture was stirred for overnight. Itwas then concentrated in vacuo to remove dioxane. To it was added water100 mL. A light yellow solid crashed out. It was isolated using aBüchner funnel, washed with cold water, dried in vacuum oven to givecompound 77.6 (230 mg, 92%). MS found for C₁₇H₁₅N₅OS as (M+H)⁺ 338.1.

Step 5: Compound 77.6 (25 mg, 0.074 mmol) was dissolved in 3 mL NMP. Toit was added MCPBA (65% pure, 22 mg, 0.081 mmol). It was stirred at RTfor 30 minutes. To it was then added cis-1,2-diaminocyclohexane (70 μL,0.60 mmol). The mixture was stirred for 30 minutes at 90° C. bath. Thismixture was then subjected to preparative HPLC to isolate the racemictitle compound 77. MS found for C₂₂H₂₅N₇O as (M+H)⁺ 404.2. UV λ=238, 337nm.

Example 62-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 5 with pyridine-3-boronic acid to replacepyridine-4-boronic acid 77.3. MS found for C₂₂H₂₅N₇O as (M+H)⁺ 404.2. UVλ=239, 311 nm.

Example 72-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridin-2-yl)phenylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 1 with 4-(pyridin-2-yl)aniline to replaceaniline 72.4. MS found for C₂₂H₂₅N₇O as (M+H)⁺ 404.2. UV λ=241, 330 nm.

Example 82-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyrimidin-5-yl)phenylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 5 with pyrimidine-5-boronic acid to replacepyridine-4-boronic acid 77.3. MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.2. UVλ=243, 308 nm. NMR (CD₃OD): δ 9.13 (s, 1H), 9.08 (s, 2H), 8.55 (s, 1H),7.82 (m, 4H), 4.44 (m, 1H), 3.74 (m, 1H), 1.94-1.60 (m, 8H) ppm.

Example 92-((1R,2S)-2-aminocyclohexylamino)-4-(4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Compound 81.1 was prepared using the same synthetic schemedemonstrated in Example 5 for 77.3 with 4-(thiazol-4-yl)aniline toreplace aniline 77.1. MS found for C₁₅H₁₃N₅OS₂ as (M+H)⁺ 344.1.

Step 2: Compound 81.1 (100 mg, 0.29 mmol) was dissolved in 4 mL NMP. Toit was added MCPBA (65% pure, 93 mg, 0.35 mmol). It was stirred at RTfor 45 minutes. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 2 mL, 0.60 mmol) andDIEA (156 μL, 0.90 mmol). The mixture was stirred for 1 hour at 90° C.bath. This mixture was diluted with ethyl acetate, washed with saturatedNaHCO₃ aqueous solution twice and water. The organic phase was driedover MgSO₄ and concentrated in vacuo to afford crude compound 81.3. MSfound for C₂₅H₃₁N₇O₃S as (M+H)⁺ 510.2.

Step 3: Compound 81.3 was stirred in a 1:1 mixture of TFA anddichloromethane at RT for 10 minutes. It was concentrated in vacuo andsubjected to reverse phase preparative HPLC to isolate the titlecompound. MS found for C₂₀H₂₃N₇OS as (M+H)⁺ 410.2. UV λ=239, 313 nm. NMR(CD₃OD): δ 9.07 (d, J=2.0 Hz, 1H), 8.53 (s, 1H), 8.01 (d, J=8.8 Hz, 2H),7.91 (s, 1H), 7.71 (d, J=7.2 Hz, 2H), 4.40 (m, 1H), 3.74 (m, 1H),1.94-1.59 (m, 8H) ppm.

Example 104-(4-(1,2,3-thiadiazol-4-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9 with4-(1,2,3-thiadiazol-4-yl)aniline to replace aniline 74.1. MS found forC₁₉H₂₂N₈OS as (M+H)⁺ 411.2. UV λ=233, 308 nm. NMR (CD₃OD): δ 9.24 (s,1H), 8.56 (s, 1H), 8.15 (d, J=5.6 Hz, 2H), 7.84 (m, 2H), 4.43 (m, 1H),3.76 (m, 1H), 1.91-1.60 (m, 8H) ppm.

Example 112-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 5 with pyridine-3-boronic acid to replacepyridine-4-boronic acid 77.3. MS found for C₂₂H₂₅N₇O as (M+H)⁺ 404.2. UVλ=240, 311 nm. NMR (CD₃OD) δ 9.05 (d, J=2.0 Hz, 1H), 8.70 (dd, J=5.6,1.2 Hz, 1H), 8.61 (d, J=6.8 Hz, 1H), 8.56 (s, 1H), 7.93-7.81 (m, 5H),4.45 (m, 1H), 3.75 (m, 1H), 1.92-1.59 (m, 8H) ppm.

Example 122-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyrimidin-5-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Compound 83.1 was prepared using the same synthetic schemedemonstrated in Example 1 for 72.7 with 3-(pyrimidin-5-yl)aniline toreplace aniline 72.4. MS found for C₂₁H₁₅N₉O₂ as (M+H)⁺ 426.1.

Step 2: Compound 83.1 (150 mg, 0.35 mmol) was dissolved in 5 mL NMP. Toit were added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 2.3 mL, 0.70 mmol) andDIEA (185 μL, 1.06 mmol). The mixture was stirred for 40 minutes at 90°C. bath. This mixture was diluted with ethyl acetate, washed with brinethree times. The organic phase was dried over MgSO₄ and concentrated invacuo to afford crude compound 83.2. MS found for C₂₆H₃₂N₈O₃ as (M+H)⁺505.2.

Step 3: Compound 83.2 was stirred in a 1:1 mixture of TFA anddichloromethane at RT for 15 minutes. It was concentrated in vacuo andsubjected to reverse phase preparative HPLC to isolate the titlecompound. MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.2. UV λ=245 nm. NMR(CD₃OD): δ 9.17 (s, 1H), 9.10 (s, 2H), 8.55 (s, 1H), 8.11 (m, 1H), 7.70(m, 1H), 7.58 (m, 2H), 4.37 (m, 1H), 3.61 (m, 1H), 1.91-1.50 (m, 8H)ppm.

Example 132-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyridin-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 5 and Example 9 with pyridine-3-boronic acid toreplace pyridine-4-boronic acid 77.3. MS found for C₂₂H₂₅N₇O as (M+H)⁺404.2. UV λ=240, 312 nm.

Example 142-((1R,2S)-2-aminocyclohexylamino)-4-(4-(thiazol-5-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Iodobenzene 77.2 (400 mg, 0.96 mmol) was dissolved in 10 mLtoluene. To it were added 5-tributylstannyithiazole (430 mg, 1.15 mmol)and Pd(PPh₃)₄ (115 mg, 0.1 mmol). The mixture was degassed using argonstream for 3 minutes and refluxed under an argon atmosphere for 1 hour.It was concentrated in vacuo and subjected to silica flash column toisolate compound 85.2 (160 mg, 45%). MS found for C₁₇H₁₆N₄O₂S₂ as (M+H)⁺373.1.

Step 2: Ethyl ester 85.2 (160 mg, 0.43 mmol) was dissolved in 30 mL THF.To it were added lithium hydroxide hydrate (55 mg, 1.3 mmol) and 5 mLwater. The mixture was stirred for 2 hours at RT. It was concentrated invacuo to remove THF and carefully treated with 1N HCl till pH reaching3. A light yellow solid crashed out from the solution. It was isolatedusing a Büchner funnel, washed with cold water, dried in vacuum oven togive compound 85.3 (120 mg, 81%). MS found for C₁₅H₁₂N₄O₂S₂ as (M+H)⁺345.1.

Step 3: Carboxylic acid 85.3 (100 mg, 0.29 mmol) was dissolved in 10 mLDMF. To it were added EDC hydrochloride (86 mg, 0.45 mmol) and HOBthydrate (61 mg, 0.45 mmol). The mixture was stirred at RT for 1 hour. Toit was then added ammonia (commercial 0.5N solution in dioxane, 3 mL,1.5 mmol). The mixture was stirred for overnight. It was thenconcentrated in vacuo to remove dioxane. To it was added water 100 mL. Alight yellow solid crashed out. It was isolated using a Büchner funnel,washed with cold water, dried in vacuum oven to give compound 85.4 (75mg, 76%). MS found for C₁₅H₁₃N₅OS₂ as (M+H)⁺ 344.1.

Step 4: Compound 85.4 (75 mg, 0.22 mmol) was dissolved in 5 mL NMP. Toit was added MCPBA (65% pure, 64 mg, 0.24 mmol). It was stirred at RTfor 30 minutes. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 1.5 mL, 0.45 mmol) andDIEA (115 μL, 0.66 mmol). The mixture was stirred for 90 minutes at 90°C. bath. This mixture was diluted with ethyl acetate, washed withsaturated NaHCO₃ aqueous solution twice and water. The organic phase wasdried over MgSO₄ and concentrated in vacuo to afford crude compound85.5. MS found for C₂₅H₃₁N₇O₃S as (M+H)⁺ 510.2.

Step 5: Compound 85.5 was stirred in a 1:1 mixture of TFA anddichloromethane at RT for 30 minutes. It was concentrated in vacuo andsubjected to reverse phase preparative

HPLC to isolate the title compound. MS found for C₂₀H₂₃N₇OS as (M+H)⁺410.2. UV λ=240, 318 nm.

Example 152-((1R,2S)-2-aminocyclohexylamino)-4-(4-(thiazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 14 with 2-tributylstannylthiazole to replace5-tributylstannylthiazole 85.1. MS found for C₂₀H₂₃N₇OS as (M+H)⁺ 410.2.UV λ=243, 332 nm. NMR (CD₃OD): δ 8.53 (s, 1H), 8.00 (d, J=8.0 Hz, 2H),7.86 (d, J=3.2 Hz, 1H), 7.77 (d, J=8.0 Hz, 2H), 7.61 (d, J=3.2 Hz, 1H),4.42 (m, 1H), 3.74 (m, 1H), 1.92-1.60 (m, 8H) ppm.

Example 164-(4-(1H-pyrazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9. MS found for C₂₀H₂₄N₈O as(M+H)⁺ 393.2. UV λ=240, 302 nm. NMR (CD₃OD): δ 8.41 (s, 1H), 8.12 (m,1H), 7.70 (d, J=8.4 Hz, 2H), 7.63 (d, J=7.2 Hz, 2H), 7.62 (s, 1H), 6.43(dd, J=2.4, 2.0 Hz, 1H), 4.27 (m, 1H), 3.60 (m, 1H), 1.83-1.45 (m, 8H)ppm.

Example 172-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9 with 3-(pyridin-3-yl)aniline toreplace aniline 74.1. MS found for C₂₂H₂₅N₇O as (M+H)⁺ 404.2. UV λ=248nm. NMR (CD₃OD): δ 9.05 (s, 1H), 8.73 (m, 1H), 8.59 (m, 1H), 8.54 (s,1H), 8.00 (broad s, 1H), 7.92 (m, 1H), 7.80 (m, 1H), 7.63-7.61 (m, 2H),4.33 (m, 1H), 3.62 (m, 1H), 1.91-1.47 (m, 8H) ppm.

Example 182-((1R,2S)-2-aminocyclohexylamino)-4-(4-morpholinophenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 5 and Example 9 with 4-morpholinoaniline toreplace aniline 74.1. MS found for C₂₁H₂₉N₇O₂ as (M+H)⁺ 412.2. UV λ=243,294 nm. NMR (CD₃OD): δ 8.45 (s, 1H), 7.49 (d, J=8.4 Hz, 2H), 7.04 (d,J=9.2 Hz, 2H), 4.30 (m, 1H), 3.85 (m, 4H), 3.71 (m, 1H), 3.19 (m, 4H),1.88-1.56 (m, 8H) ppm.

Example 192-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyrimidin-5-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 5 and Example 9 with pyrimidine-5-boronic acidto replace pyridine-4-boronic acid 77.3. MS found for C₂₁H₂₄N₈O as(M+H)⁺ 405.2. UV λ=242, 307 nm. NMR (CD₃OD): δ 9.13 (s, 1H), 9.08 (s,2H), 8.55 (s, 1H), 7.82 (m, 4H), 4.44 (m, 1H), 3.74 (m, 1H), 1.95-1.58(m, 8H) ppm.

Example 202-((1R,2S)-2-aminocyclohexylamino)-4-(4-(morpholinomethyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9 with 4-(morpholinomethyl)anilineto replace aniline 74.1. MS found for C₂₂H₃₁N₇O₂ as (M+H)⁺ 426.3. UVλ=244, 293 nm.

Example 212-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridazin-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 14 with 4-tributylstannylpyridazine to replace5-tributylstannylthiazole 85.1. MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.3.UV λ=239, 327 nm. NMR (CD₃OD): δ 9.59 (m, 1H), 9.21 (d, J=9.2 Hz, 1H),8.58 (s, 1H), 8.08 (m, 1H), 7.96-7.85 (m, 5H), 4.47 (m, 1H), 3.77 (m,1H), 1.93-1.59 (m, 8H) ppm.

Example 222-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyrazin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 14 with 2-tributylstannylpyrazine to replace5-tributylstannylthiazole 85.1. MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.3.UV λ=235, 319 nm. NMR (CD₃OD): δ 9.11 (s, 1H), 8.66 (broad s, 1H), 8.56(s, 1H), 8.51 (broad s, 1H), 8.14 (m, 2H), 7.85 (m, 2H), 4.44 (m, 1H),3.77 (m, 1H), 1.92-1.58 (m, 8H) ppm.

Example 232-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyridin-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 5 and Example 9. MS found for C₂₂H₂₅N₇O as(M+H)⁺ 404.2. UV λ=239, 334 nm. NMR (CD₃OD): δ 8.76 (d, J=6.8 Hz, 2H),8.60 (s, 1H), 8.29 (d, J=6.4 Hz, 2H), 8.05-7.96 (m, 4H), 4.49 (m, 1H),3.76 (m, 1H), 1.93-1.60 (m, 8H) ppm.

Example 244-(4-(1H-1,2,3-triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

Step 1: Compound 72.3 (1.18 g, 5.3 mmol) was dissolved in 40 mL NMP andstirred at RT. To it were added 4-azidoaniline hydrochloride 95.1 (1.00g, 5.9 mmol) and then DIEA (2.21 mL, 12.7 mmol) dropwise using syringe.The mixture was stirred for 1 hour and diluted with ethyl acetate. Itwas washed with brine four times, dried and concentrated in vacuo toafford compound 95.2 (1.59 g, 94%). MS found for C₁₃H₁₁ClN₆O₂ as (M+H)⁺319.2.

Step 2: Ethyl ester 95.2 (1.59 g, 5.0 mmol) was dissolved in 50 mL THF.To it were added lithium hydroxide hydrate (420 mg, 10 mmol) and 5 mLwater. The mixture was stirred for 4 hours and to it was carefully added3N HCl solution till pH reaching 3. The mixture was concentrated invacuo to remove THF. The residue was taken into ethyl acetate and washedwith brine twice. It was dried and concentrated in vacuo to givecompound 95.3 (1.58 g, 99%) as a solid. MS found for C₁₁H₇ClN₆O₂ as(M+H)⁺ 291.2.

Step 3: Carboxylic acid 95.3 (1.58 g, 5.0 mmol) was dissolved in 40 mLDMF. To it were added EDC hydrochloride (1.44 g, 7.5 mmol) and HOBthydrate (1.02 g, 7.5 mmol). The mixture was stirred at RT for 50minutes. To it was then added ammonia (commercial 0.5N solution indioxane, 50 mL, 25 mmol). The mixture was stirred for 7 hours. It wasthen concentrated in vacuo and a solid crashed out. It was collected,washed and dried in vacuum oven to afford compound 95.4 (1.30 g, 67%).MS found for C₁₇H₁₂N₁₀O₂ as (M+H)⁺ 389.3.

Step 4: Compound 95.4 (300 mg, 0.77 mmol) was dissolved in 20 mL NMP. Toit were added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 5.1 mL, 1.5 mmol) andDIEA (400 μL, 2.3 mmol). The mixture was stirred for 90 minutes at 90°C. bath. This mixture was diluted with ethyl acetate, washed with brinethree times. The organic phase was dried over MgSO₄, concentrated invacuo and subjected to flash column to afford compound 95.5 (420 mg,90%). MS found for C₂₂H₂₉N₉O₃ as (M+H)⁺ 467.3.

Step 5: Compound 95.5 (420 mg, 0.70 mmol) was stirred in 10 mL methanol.To it were added trimethylsilylacetylene (200 mg, 1.4 mmol), CuI (400mg, 2.1 mmol) and DBU (313 μL, 2.1 mmol). The mixture was stirred at RTfor 4 hours. It was diluted with ethyl acetate, washed with saturatedammonium chloride aqueous solution and brine twice. The organic phasewas dried, filtered and concentrated in vacuo to afford crude compound95.6. MS found for C₂₄H₃₁N₉O₃ as (M+H)⁺ 494.4.

Step 6: Crude compound 95.6 was stirred in a 1:1 mixture of TFA anddichloromethane at RT for 90 minutes. It was concentrated in vacuo andsubjected to reverse phase preparative HPLC to isolate the titlecompound. MS found for C₁₉H₂₃N₉₀ as (M+H)⁺ 394.4. UV λ=242, 300 nm.

Example 252-((1R,2S)-2-aminocyclohexylamino)-4-(4-(1-methyl-1H-imidazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 14 with 1-methyl-2-tributylstannylimidazole toreplace 5-tributylstannylthiazole 85.1. MS found for C₂₁H₂₆N₈O as (M+H)⁺407.4. UV λ=241, 300 nm. NMR (CD₃OD): δ 8.97 (s, 1H), 8.59 (s, 1H), 7.90(m, 2H), 7.64 (m, 3H), 4.46 (m, 1H), 3.91 (s, 3H), 3.73 (m, 1H),1.92-1.58 (m, 8H) ppm.

Example 262-((1R,2S)-2-aminocyclohexylamino)-4-(4-(2-oxopyridin-1(2H)-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Iodobenzene 77.2 (440 mg, 1.06 mmol) was dissolved in 10 mLDMSO. To it were added 2-hydroxypyridine 97.1 (202 mg, 2.12 mmol), K₂CO₃(293 mg, 2.12 mmol), CuI (61 mg, 0.32 mmol) and 8-hydroxyquinoline (46mg, 0.32 mmol). The mixture was stirred in 120° C. bath for 5 hours. Tothe mixture were added lithium hydroxide hydrate (126 mg, 3 mmol) and 10mL water. The mixture was stirred for overnight. To it was carefullyadded 1N HCl till pH reaching 3. The mixture was filtered through celiteand subjected to reverse phase prep HPLC to isolate compound 97.2 (75mg, 20%). MS found for C₁₇H₁₄N₄O₃S as (M+H)⁺ 354.3.

Step 2: Carboxylic acid 97.2 (75 mg, 0.21 mmol) was dissolved in 10 mLDMF. To it were added EDC hydrochloride (61 mg, 0.32 mmol) and HOBthydrate (44 mg, 0.32 mmol). The mixture was stirred at RT for 90minutes. To it was then added ammonia (commercial 0.5N solution indioxane, 2 mL, 1 mmol). The mixture was stirred for 2 hours. It was thenconcentrated in vacuo and subjected to reverse phase preparative HPLC toisolate compound 97.3 (40 mg, 53%). MS found for C₁₇H₁₅N₅O₂S as (M+H)⁺355.3.

Step 3: Compound 97.3 (40 mg, 0.11 mmol) was dissolved in 5 mL NMP. Toit was added MCPBA (65% pure, 37 mg, 0.14 mmol). It was stirred at RTfor 1 hour. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 0.73 mL, 0.22 mmol) andDIEA (115 μL, 0.66 mmol). The mixture was stirred for 1 hour at 90° C.bath. This mixture was diluted with ethyl acetate, washed with saturatedNaHCO₃ aqueous solution twice and water. The organic phase was driedover MgSO₄ and concentrated in vacuo to afford crude compound 97.3. MSfound for C₂₇H₃₃N₇O₄ as (M+H)⁺ 520.4.

Step 4: Compound 97.4 was stirred in a 4:1 mixture of TFA anddichloromethane at 50° for 1 hour. It was concentrated in vacuo andsubjected to reverse phase preparative HPLC to isolate the titlecompound. MS found for C₂₂H₂₅N₇O₂ as (M+H)⁺ 420.4. UV λ=241, 296 nm.

Example 274-(4-(1H-imidazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 and Example 9 with 4-(1H-imidazol-1-yl)anilineto replace aniline 74.1. MS found for C₂₀H₂₄N₈O as (M+H)⁺ 393.4. UVλ=246, 292 nm. NMR (CD₃OD): δ 9.41 (s, 1H), 8.59 (s, 1H), 8.05-7.95 (m,3H), 7.79-7.76 (m, 3H), 4.46 (m, 1H), 3.71 (m, 1H), 1.93-1.59 (m, 8H)ppm.

Example 282-((1R,2S)-2-aminocyclohexylamino)-4-(4-(piperidin-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9 with 4-(piperidin-1-yl)anilineto replace aniline 72.4. MS found for C₂₂H₃₁N₇O as (M+H)⁺ 410.4. UVλ=246, 287 nm. NMR (CD₃OD): δ 8.55 (s, 1H), 7.81 (m, 2H), 7.59 (m, 2H),4.41 (m, 1H), 3.71 (m, 1H), 3.58 (m, 4H), 2.02 (m, 4H), 1.90-1.80 (m,8H), 1.60 (m, 2H) ppm.

Example 292-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-morpholinophenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 1 with3-fluoro-4-morpholinoaniline to replace aniline 72.4. MS found forC₂₁H₂₈FN₇O₂ as (M+H)⁺ 430.4. UV λ=239, 309 nm. NMR (CD₃OD): δ 8.49 (s,1H), 7.63 (m, 1H), 7.22 (m, 1H), 7.08 (dd, J=9.6, 8.8 Hz, 1H), 4.34 (m,1H), 3.85 (m, 4H), 3.76 (m, 1H), 3.08 (m, 4H), 1.92-1.58 (m, 8H) ppm.

Example 302-((1R,2S)-2-aminocyclohexylamino)-4-(4-(3-(trifluoromethyl)-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 and Example 9 with4-(3-(trifluoromethyl)-1H-pyrazol-1-yl)aniline to replace aniline 74.1.MS found for C₂₁H₂₃F₃N₈O as (M+H)⁺ 461.4. UV λ=241, 236 nm. NMR (CD₃OD):δ 8.54 (s, 1H), 8.38 (broad s, 1H), 7.85-7.80 (m, 4H), 6.84 (d, J=2.0Hz, 1H), 4.41 (m, 1H), 3.71 (m, 1H), 1.91-1.59 (m, 8H) ppm.

Example 314-(3-(1H-pyrazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 and Example 9 with 3-(1H-pyrazol-1-yl)anilineto replace aniline 74.1. MS found for C₂₀H₂₄N₈O as (M+H)⁺ 393.4. UVλ=247 nm.

Example 322-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

A mixture of 3,4-difluoronitrobenzene (1.00 mL, 9.03 mmol), pyrazole(0.615 g, 9.04 mmol) and K2CO3 (2.50 g, 18.1 mmol) in DMF (10 mL) wasstirred at room temperature overnight. Water (30 mL) was added to induceprecipitation. The precipitate was collected, dried on vacuum togivel-(2-fluoro-4-nitrophenyl)-1H-pyrazole as a solid (1.80 g). MS 208.3(M+H)

A suspension of 1-(2-fluoro-4-nitrophenyl)-1H-pyrazole (1.80 g, 8.70mmol) and Pd—C (10%, 0.200 g) in MeOH (20 mL) (containing 10 drops of 6NHCl) was hydrogenated under balloon H2 overnight. The mixture wasfiltered through celite. The filtrate was concentrated in vacuo. Theresidue was dried on vacuum to give3-fluoro-4-(1H-pyrazol-1-yl)benzenamine as a solid (1.55 g). MS 178.3(M+H)

To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (280 mg,1.27 mmol) and 3-fluoro-4-(1H-pyrazol-1-yl)benzenamine (230 mg, 1.30mmol) in CH₃CN (8 mL) at room temperature, DIEA (0.442 mL, 2.54 mmol)was added. The mixture was stirred at room temperature for 48 h. Water(15 mL) was added to induce precipitation. The precipitate wascollected, dried on vacuum to give ethyl2-chloro-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxylateas a solid (275 mg). MS 362.3 and 364.3 (M+H, Cl pattern)

To a solution of ethyl2-chloro-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxylate(275 mg, 0.761 mmol) in THF (4 mL), aq. 1N LiOH (1.25 mL, 1.25 mmol) wasadded. The mixture was stirred at room temperature overnight. Uponacidification of the mixture with 1N HCl, white solids precipitated out,which were collected, and dried on vacuum to give2-chloro-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxylicacid (230 mg). MS 334.3 and 336.3 (M+H, Cl pattern)

To a solution of2-chloro-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxylicacid (230 mg, 0.690 mmol) and HOBt (158 mg, 1.03 mmol) in DMF (4 mL),EDC (200 mg, 1.04 mmol) was added. The mixture was stirred at roomtemperature for 1.5 h. Ammonia (0.5 M in dioxane, 6.00 mL, 3.00 mmol)was added. It was stirred at room temperature overnight. Water and EtOAcwere added. The organic phase was separated, washed with 1 N HCl, thenwith 5% NaHCO₃, dried over Na₂SO₄, concentrated in vacuo to give2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide(184 mg). MS 432.4 (M+H)

A solution of tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (0.30 M inNMP, 2.00 mL, 0.600 mmol) in NMP (2 mL) was added to2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide(184 mg, 0.427 mmol). DIEA (0.150 mL, 0.863 mmol) was also added. Themixture was stirred at 90 C overnight. After being cooled to roomtemperature, water was added to induce precipitation.

The precipitate was collected, then dissolved in CH₂Cl₂ (5 mL) and TFA(4 mL). The solution was stirred at room temperature for 30 min. It wasconcentrated in vacuo. The residue was purified by HPLC to give thetitled compound (103 mg). MS 411.5 (M+H). UV λ=238.8, 304.8 nm.

Example 332-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

A mixture of 2,5-diiodofluorobenzene (1.36 g, 3.90 mmol),tert-butylcarbamate (0.454 g, 3.88 mmol), xantphos (0.233 g, 0.40 mmol)and Cs₂CO₃ (dry powder, 2.48 g, 7.61 mmol) in THF (12 mL) was degassedwith Ar before being charged with Pd₂(dba)₃ (0.071 g, 0.078 mmol). Itwas stirred at 75 C overnight. After being cooled to room temperature,water and EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was loaded to a flash column,eluted with 0-10% EtOAc in hexane to give tert-butyl3-fluoro-4-iodophenylcarbamate (716 mg).

A solution of tert-butyl 3-fluoro-4-iodophenylcarbamate (500 mg, 1.48mmol) and 4-(tributylstannyl)thiazole (0.578 mL, 1.80 mmol) in dioxane(6 mL) was degassed with Ar before being charged with Pd(Ph3P)₄ (170 mg,0.147 mmol). It was stirred at 100 C for 34 h, then concentrated invacuo. The residue was loaded to a flash column, eluted with 0-25% EtOAcin hexane to give tert-butyl 3-fluoro-4-(thiazol-4-yl)phenylcarbamate(270 mg). MS 295.3 (M+H)

A solution of tert-butyl 3-fluoro-4-(thiazol-4-yl)phenylcarbamate (270mg, 0.92 mmol) in CH₂Cl₂ (2 mL) and TFA (4 mL) was stirred at roomtemperature for 60 min. It was then concentrated in vacuo. The residuewas dissolved in CH₂Cl₂ (20 mL), which was washed with 5% NaHCO₃, driedover Na₂SO₄, concentrated in vacuo to give3-fluoro-4-(thiazol-4-yl)benzenamine (145 mg) as free base. MS195.2(M+H)

To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (165 mg,0.746 mmol) and 3-fluoro-4-(thiazol-4-yl)benzenamine (145 mg, 0.747mmol) in CH₃CN (5 mL) at room temperature, DIEA (0.260 mL, 1.49 mmol)was added. The mixture was stirred at room temperature overnight, duringwhich time solids precipitated out. The precipitate was collected, driedon vacuum to give ethyl2-chloro-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxylateas a solid (186 mg). MS 379.3 and 381.3 (M+H, Cl pattern)

To a suspension of ethyl2-chloro-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxylate(186 mg, 0.491 mmol) in THF (4 mL), aq. 1N LiOH (1.00 mL, 1.00 mmol) wasadded. The suspension became clear with stirring. The mixture was thenstirred at room temperature overnight. Upon acidification of the mixturewith 1N HCl, white solids precipitated out, which were collected, anddried on vacuum to give2-chloro-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxylicacid (158 mg). MS 351.2 and 353.3 (M+H, Cl pattern)

To a solution of2-chloro-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxylicacid (158 mg, 0.450 mmol) and HOBt (103 mg, 0.673 mmol) in DMF (4 mL),EDC (130 mg, 0.678 mmol) was added. The mixture was stirred at roomtemperature for 1 h. Ammonia (0.5 M in dioxane, 4.50 mL, 2.25 mmol) wasadded. It was stirred at room temperature overnight. Water and EtOAcwere added. The organic phase was separated, washed with 5% NaHCO₃,dried over Na₂SO₄, concentrated in vacuo. The residue was dissolved inH₂O and CH₃CN (50:50). The insoluble was collected and dried on vacuumto give2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamideas a solid (36 mg). MS 449.4 (M+H)

A solution of tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (0.30 M inNMP, 1.00 mL, 0.300 mmol) in NMP (1 mL) was added to2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide(12 mg, 0.027 mmol). DIEA (0.050 mL, 0.29 mmol) was also added. Themixture was stirred at 90 C for 1 h. After being cooled to roomtemperature, TFA (1 mL) was added. The solution was stirred at roomtemperature for 60 min. The mixture was purified by HPLC to give thetitled compound (5 mg). MS 428.4 (M+H). UV λ=229.8, 313.8 nm.

Example 342-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide

A mixture of 3,4-difluoronitrobenzene (1.00 mL, 9.03 mmol), imidazole(0.615 g, 9.04 mmol) and K2CO3 (2.50 g, 18.1 mmol) in DMF (10 mL) wasstirred at room temperature overnight. Water (20 mL) was added to induceprecipitation. The precipitate was collected, dried on vacuum to give1-(2-fluoro-4-nitrophenyl)-1H-imidazole as a solid (1.81 g). MS 208.2(M+H)

A mixture of 1-(2-fluoro-4-nitrophenyl)-1H-imidazole (1.81 g, 8.74 mmol)and Pd—C (10%, 0.200 g) in MeOH (20 mL) (containing 10 drops of 6N HCl)was hydrogenated under balloon H2 overnight. The mixture was filteredthrough celite. The filtrate was concentrated in vacuo. The residue wasdried on vacuum to give 3-fluoro-4-(1H-imidazol-1-yl)benzenamine as asolid (1.37 g). MS178.3 (M+H)

To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (280 mg,1.27 mmol) and 3-fluoro-4-(1H-imidazol-1-yl)benzenamine (230 mg, 1.30mmol) in CH₃CN (8 mL) at room temperature, DIEA (0.442 mL, 2.54 mmol)was added. The mixture was stirred at room temperature overnight. Waterand EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo to give ethyl2-chloro-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxylateas a solid (383 mg). MS 362.4 and 364.3 (M+H, Cl pattern)

To a solution of ethyl2-chloro-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxylate(383 mg, 1.06 mmol) in THF (5 mL), aq. 1N LiOH (2.00 mL, 2.00 mmol) wasadded. The mixture was stirred at room temperature overnight. THF wasremoved in vacuo. Upon acidification with 1N HCl, the mixture waspurified by HPLC to give2-chloro-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxylicacid (68 mg). MS 334.1 and 336.1 (M+H, Cl pattern)

To a solution of2-chloro-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxylicacid (68 mg, 0.20 mmol) and HOBt (63 mg, 0.41 mmol) in DMF (2 mL), EDC(60 mg, 0.31 mmol) was added. The mixture was stirred at roomtemperature for 1.5 h. Ammonia (0.5 M in dioxane, 0.800 mL, 0.40 mmol)was added. It was stirred at room temperature overnight. More EDC (100mg, 0.52 mmol) was added. It was stirred for another 24 h. Water andEtOAc were added. The organic phase was separated, washed with 5%NaHCO₃, dried over Na₂SO₄, concentrated in vacuo to give2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide(57 mg). MS 432.2 (M+H)

A solution of tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (0.30 M inNMP, 1.00 mL, 0.300 mmol) in NMP (1 mL) was added to2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(3-fluoro-4-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide(57 mg, 0.13 mmol). DIEA (0.100 mL, 0.57 mmol) was also added. Themixture was stirred at 90 C for 1 h. After being cooled to roomtemperature, water and EtOAc were added. The organic phase wasseparated, washed with 5% NaHCO₃, dried over Na₂SO₄, concentrated invacuo. The residue was dissolved in CH₂Cl₂ (1 mL) and TFA (1 mL). Thesolution was stirred at room temperature for 60 min. It was concentratedin vacuo. The residue was purified by HPLC to give the titled compound(23 mg). MS 411.3 (M+H). UV λ=247.8, 299.8 nm.

Example 352-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(4-methyl-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The titled compound was prepared analogously according to the proceduredescribed in Example 34. MS 425.3 (M+H). UV λ=247.8, 311.8 nm.

Example 362-((1R,2S)-2-aminocyclohexylamino)-4-(2-fluoro-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

Compound 107.1 was synthesized as demonstrated in Example 1 for 72.7using 2-fluoro-4-(1H-pyrazol-1-yl)aniline to replace 72.4. To 107.1(0.054 g, 0.125 mmol) was added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 0.6 mL, 0.18 mmol) andDIEA (33 μL, 0.18 mmol). After stirred for 2 h at 65° C. bath, themixture was diluted with water, the resulting precipitates werecollected by filtration to give 107.2. To a mixture of 107.2 in DCM (0.5mL) was added TFA (0.5 mL), after stirred at room temperature for 10min, the solution was concentrated and the residue was purified bypreparative HPLC to give title compound 107. MS found for C₂₀H₂₃FN₈O as(M+H)⁺ 411.1. UV: λ=201.6, 240.4, 289.0.

Example 372-((1R,2S)-2-aminocyclohexylamino)-4-(3-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: To a mixture of Iodobenzene 108.1 (200 mg, 0.46 mmol) in toluene(2 mL) was added 5-tributylstannylthiazole 108.2 (173 mg, 0.46 mmol) andPd(PPh₃)₄ (53 mg, 0.046 mmol). The mixture was degassed using argonstream for 3 minutes and refluxed under an argon atmosphere for 2 hour.It was concentrated in vacuo and subjected to silica flash column toisolate compound 108.3 (100 mg).

Step 2: To a mixture of Ethyl ester 108.3 (84 mg, 0.22 mmol) in THF (0.8mL) was added a solution of lithium hydroxide hydrate (8 mg, 0.35 mmol)in water (0.5 mL). After stirred for 2 hours at RT, it was concentratedin vacuo to remove THF and carefully treated with 1N HCl till pHreaching 3. The resulting precipitates were collected by filtration togive compound 108.4 (74 mg).

Step 3: To a mixture of carboxylic acid 108.4 (74 mg) in DMF (1 mL) wereadded EDC hydrochloride (60 mg) and HOBt hydrate (48 mg). The mixturewas stirred at RT for 1 hour. To it was then added ammonia (commercial0.5N solution in dioxane, 1 mL). The mixture was stirred for 1 h at roomtemperature. It was then concentrated in vacuo to remove dioxane. To itwas added water, the precipitates were collected by filtration to givecompound 108.5.

Step 4: Compound 108.5 (55 mg, 0.16 mmol) was dissolved in 0.3 mL NMP.To it was added MCPBA (65% pure, 47 mg, 0.18 mmol). It was stirred at RTfor 30 minutes. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 57.2 (0.3 M, 0.8 mL, 0.24 mmol) andDIEA (57 μL, 0.32 mmol). The mixture was stirred for 60 minutes at 80°C. bath. This mixture was diluted with water, the resulting precipitateswas collected by filtration to five 108.6, which was stirred in a 1:1mixture of TFA and dichloromethane at RT for 30 minutes. It wasconcentrated in vacuo and subjected to reverse phase preparative HPLC toisolate the title compound. MS found for C₂₀H₂₃N₇OS as (M+H)⁺ 410.2. UV:λ=202.8, 245.2.

Example 382-((1R,2S)-2-aminocyclohexylamino)-4-(2-fluoro-4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 14. MSfound for C₂₀H₂₂FN₇OS as (M+H)⁺ 428.5. UV: λ=240.4, 312.8

Example 392-((1R,2S)-2-aminocyclohexylamino)-4-(3-(thiazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 37 using2-tributylstannylthiazole to replace 5-tributylstannylthiazole 108.2. MSfound for C₂₀H₂₃N₇OS as (M+H)⁺ 410.2 UV: λ=206.3, 242.8, 291.4

Example 402-((1R,2S)-2-aminocyclohexylamino)-4-(3-(thiazol-5-yl)phenylamino)pyrimidine-5-carboxamide

The title compound III was synthesized as demonstrated in Example 37using 4-tributylstannylthiazole to replace 5-tributylstannylthiazole108.2. MS found for C₂₀H₂₃N₇OS as (M+H)⁺ 410.5 UV: λ=201.6, 244.0, 277.1

Example 412-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyridin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 37 using2-tributylstannylpyridine to replace 5-tributylstannylthiazole 108.2. MSfound for C₂₂H₂₅N₇O as (M+H)⁺ 404.2; UV: λ=242.8, 292.6

Example 422-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyrazin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 37 using2-tributylstannylpyrazine to replace 5-tributylstannylthiazole 108.2. MSfound for C₂₁H₂₄N₈O as (M+H)⁺ 405.3 UV: λ=244.0, 289.0

Example 432-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyridazin-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 37 using4-tributylstannylpyridazine to replace 5-tributylstannylthiazole 108.2.MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.2 UV: λ=246.3

Example 442-((1R,2S)-2-aminocyclohexylamino)-4-(3-(1-methyl-1H-imidazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 37 using1-methyl-2-tributylstannyl-1H-imidazole to replace5-tributylstannylthiazole 108.2. MS found for C₂₁H₂₆N₈O as (M+H)⁺ 407.2.UV: λ=242.8

Example 452-((1R,2S)-2-aminocyclohexylamino)-4-(3-(6-methoxypyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: To a mixture of Iodobenzene 108.1 (300 mg, 0.72 mmol) inp-dioxane (5 mL) were added 6-methoxypridin-3-ylboronic acid 116.1 (121mg, 0.80 mmol) and 1M Na₂CO₃ (21. mL) followed by PdCl2(PPh₃)₂ (51 mg,0.07 mmol). The mixture was degassed using argon stream for 3 minutesand heated at 85° C. under an argon atmosphere for 2 hour. It wasdiluted with DCM, the organic layer was washed with Sat. NaHCO₃, brine,dried and concentrated to give crude residue, which was purified bycolumn chromatography to give 116.2 (160 mg).

Step 2: To a mixture of Ethyl ester 116.2 (160 mg) in THF (1.6 mL) wasadded a solution of lithium hydroxide hydrate (20 mg) in water (1 mL).After stirred for 24 hours at RT, it was concentrated in vacuo to removeTHF and carefully treated with 1N HCl till pH reaching 3. The resultingprecipitates were collected by filtration to give compound 116.3.

Step 3: To a mixture of carboxylic acid 116.3 in DMF (1.8 mL) were addedEDC hydrochloride (193 mg) and HOBt hydrate (153 mg). The mixture wasstirred at RT for 10 min. To it was then added ammonia (commercial 0.5Nsolution in dioxane, 2 mL). The mixture was stirred for 1 h at roomtemperature. It was then concentrated in vacuo to remove dioxane. To itwas added water, the precipitates were collected by filtration to givecompound 116.4.

Step 4: Compound 116.4 (80 mg, 0.22 mmol) was dissolved in 0.5 mL NMP.To it was added MCPBA (65% pure, 64 mg, 0.24 mmol). It was stirred at RTfor 30 minutes. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 1.2 mL, 0.3 mmol) andDIEA (78 μL, 0.44 mmol). The mixture was stirred for 60 minutes at 85°C. bath. This mixture was diluted with water, the resulting precipitateswas collected by filtration to give 116.5, which was stirred in a 1:1mixture of TFA and dichloromethane at RT for 30 minutes. It wasconcentrated in vacuo and subjected to reverse phase preparative HPLC toisolate the title compound. MS found for C₂₃H₂₇N₇O₂ as (M+H)⁺ 434.3 UV:λ=247.5

Example 462-((1R,2S)-2-aminocyclohexylamino)-4-(4-(6-methoxypyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was synthesized as demonstrated in Example 45 usingethyl-4-(4-iodophenylamino)-2-(methylthio)pyrimidine-5-carboxylate toreplace 108.1. MS found for C₂₃H₂₇N₇O₂ as (M+H)⁺ 434.3. UV: λ=240.4,306.8

Example 472-((1R,2S)-2-aminocyclohexylamino)-4-(3-methyl-4-morpholinophenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 and Example 9 with3-methyl-4-morpholinoaniline to replace aniline 72.4. MS found forC₂₂H₃₁N₇O₂ as (M+H)⁺ 426.3. UV: λ=240.4, 296.1

Example 482-((1R,2S)-2-aminocyclohexylamino)-4-(4-(2-aminopyridin-4-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: A mixture of 4-Nitro-phenyl-Boronic acid (159 mgs, 0.95 mmol),N,N-Bis(tert-butyloxycarbonyl)amino-4-Iodo pyridine (U.S. Pat. No.6,831,175) (200 mgs, 0.47 mmol), potassium carbonate (329 mgs, 2.38mmol), Tetrakis(triphenylphosphine)-palladium(0) (220 mgs, 0.19 mmol),DME (5 mL), water (0.6 mL) was heated in microwave (Emry's Optimizer) at120° C. for 15 min. The reaction was repeated four more times. Thecombined reactions were poured into ethyl acetate and washed with water(2×), brine (1×) and concentrated. The concentrate was then purified byflash column chromatography on silica gel (3:7 Ethyl acetate/Hexanes) toprovide N,N-Bis(tert-butyloxycarbonyl)amino-(4-nitrophenyl)pyridine119.1. MS found for C₂₁H₂₅N₃O₆ as (M+H)⁺ 416.0.

Step 2: Compound 119.1 (955 mgs, 2.3 mmol) was dissolved in ethanol (40mL) with few drops of acetic acid and hydrogenated at 1 atmosphere ofhydrogen in the presence of 10% Pd/C (wet) for 5 h. The reaction wasfiltered, concentrated to give 119.2 (782 mgs, 88%) as dark brown solid.MS found for C₂₁H₂₇N₃O₄ as (M+H)⁺ 386.0.

Step 3: To a solution of Dichloropyrimidine 119.3 (180 mgs, 0.816 mmol)in acetonitrile (3 mL) was added a suspension of 119.2 (314 mgs, 0.816mmol), diisopropylamine (0.16 mL, 0.897 mmol) in acetonitrile (8 mL) at0° C. Reaction mixture was then slowly warmed to rt and stirredovernight. The reaction mixture was then diluted with water andextracted with ethylacetate (2×). The combined ethyl acetate layers werewashed with brine, dried (MgSO₄), filtered and concentrated underreduced pressure to yield 119.4. MS found for C₂₈H₃₂ClN₅O₆ as (M+H)⁺570.0.

Step 4: Crude ethyl ester 119.4 (460 mgs, 0.81 mmol) was diluted with1,4-dioxane (5 mL) followed by aqueous lithium hydroxide (1.0 M, 0.8 mL,0.8 mmol) and stirred at rt until all starting material had beenconverted to the carboxylic acid. The reaction was then diluted withwater (30 mL) and acidified with 1N HCl (1.0 mL). The resultingsuspension was then filtered, washed with water and dried giving 385 mgsof the carboxylic acid 119.5 (88%). MS found for C₂₆H₂₈ClN₅O₆ as (M+H)⁺542.0.

Step 5: To carboxylic acid 119.5 (385 mgs, 0.71 mmol), EDC (204 mgs,1.06 mmol), HOBt (163 mgs, 1.06 mmol) in N,N-dimethylformamide (3.6 mL)was added ammonia (0.5 M in 1,4-dioxane, 3.6 mL, 1.8 mmol) and stirredovernight. The reaction mixture was then diluted with water and theprecipitate collected by filtration affording the desired product 119.6(394 mgs, 87%). MS found for C₃₂H₃₃N₉O₆ as (M+H-Boc)⁺540.0.

Step 6 and Step 7: A mixture of Benzotriazolyl ether 119.6 (90 mgs,0.140 mmol), tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (36 mgs,0.170 mmol), DIPEA (0.07 mL, 0.420 mmol) in iso-propanol (2 mL) washeated in microwave (Emry's Optimizer) at 130° C. for 20 min. Thereaction mixture was concentrated and then was treated with 4.0M HCl indioxane (6.0 mL). After 1 h at rt, concentrate the reaction mixture anddiluted with water and acetonitrile and directly purified by preparativeHPLC affording the desired product 119 (43.6 mgs, 75%) as tan solid,after lyophilization. MS found for C₂₂H₂₆F₂N₈O as (M+H)⁺ 419.5.

Example 492-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-(aminomethyl)piperidin-1-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using tert-butyl(1-(4-aminophenyl)piperidin-4-yl)methylcarbamate (prepared from4-Fluoronitrobenzene) using a procedure similar to that described inExample 48. MS found for C₂₃H₃₄N₈O as (M+H)⁺ 439.6.

Example 504-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexyl-amino)pyrimidine-5-carboxamide

Ethyl4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-chloropyrimidine-5-carboxylate121.1

To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (1.4) (600mg, 2.714 mmol), in CH₃CN (12 mL), was added DIEA (0.750 mL, 4.313 mmol,1.589 equiv), followed by 4-(1H-1,2,4-triazol-1-yl)aniline (465 mg,2.902 mmol, 1.07 equiv). The mixture was stirred at room temperature for20 h, then diluted with water (6×) to precipitate ii. The solid productwas collected by filtration, rinsed with water (100 mL), and air dried;Yield, 850 mg (91%).

4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-chloropyrimidine-5-carboxylicacid 121.2

To a solution of Ethyl4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-chloropyrimidine-5-carboxylate(ii) (870 mg, 2.523 mmol), in THF (12 mL), was added 1M LiOH (3.30 mL,3.30 mmol, 1.30 equiv), and the mixture was stirred at room temperaturefor 3 h. Then THF was rotavopped off, and the aqueous mixture wasacidified with 2N HCl to pH 2. The precipitated solid product wascollected by filtration, rinsed with water, and air dried; yield 712 mg(89%).

4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-(1H-benzo[d][1,2,3]triazol-1-yloxy)pyrimidine-5-carboxamide 121.3

To a solution of4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-chloropyrimidine-5-carboxylicacid (iii) (730 mg, 2.305 mmol) in DMF (15 mL), at room temperature, wasadded HOBt (470 mg, 3.478 mmol, 1.508 equiv), and EDC.HCl (690 mg, 3.599mmol, 1.561 equiv), and the mixture was stirred for 45 minutes. Thencooled it to 0° C., and treated with 0.5 M NH3 in dioxane (14.0 ml, 7.0mmol, 3.0 equiv), and stirred the mixture at room temperature for 18 h.Then diluted it with water (7×) to precipitate iv. Collected it byfiltration, rinsed with water, and air dried; yield 841 mg (88%). MS:415.1 (M+H), and 437.1 (M+Na).

4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)-pyrimidine-5-carboxamide121

To a solution of cis-cyclohexane-1,2-diamine (100 mg, 0.876 mmol, 6.08equiv), in CH₃CN (1.50 mL), was added4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-(1H-benzo[d][1,2,3]triazol-1-yloxy)pyrimidine-5-carboxamide(iv) (60 mg, 0.144 mmol), and the mixture was stirred for 19 h. Then itwas diluted with water (8×), and the solid product was collected byfiltration, rinsed with water, and air dried. It was purified by RP HPLCto afford it as its TFA salt; yield 59 mg (81%). MS: 394.1 (M+H), and416.1 (M+Na).

Example 512-((1R,2S)-2-aminocyclohexylamino)-4-(4-(isoxazol-5-yl)phenylamino)pyrimidine-5-carboxamide

2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(4-(isoxazol-5-yl)phenylamino)pyrimidine-5-carboxamide122.1: This compound was prepared using the procedure described for thesynthesis of 121.3 starting from ethyl2,4-dichloropyrimidine-5-carboxylate (1.4) and 4-(isoxazol-5-yl)aniline.

tert-butyl(1S,2R)-2-(5-carbamoyl-4-(4-(isoxazol-5-yl)phenylamino)pyrimidin-2-ylamino)cyclohexylcarbamate122.2

The compound 210.1 (100 mg, 0.241 mmol), in N-methylpyrrolidinone (NMP)was mixed with tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (0.3 mmol,1.244 equiv), and DIEA (0.120 mL, 0.690 mmol, 2.83 equiv), and heated to50° C. for 4 h. Then it was partitioned between EtOAc and water. Thecombined EtOAc extract was dried over anhydrous Na₂SO₄ and concentratedto yield 210.2, which was used as such for the next reaction.

2-((1R,2S)-2-aminocyclohexylamino)-4-(4-(isoxazol-5-yl)phenylamino)pyrimidine-5-carboxamide122

The crude 122.2 was diluted with CH₂Cl₂ (5 mL) and anilsole (0.700 mL,excess) was added, followed by CF₃COOH (5 mL). The mixture was stirredat room temperature for 5 h, and then concentrated to dryness. It waspurified by RP-HPLC to yield pure 210 as colorless puffs, 101 mg (83%).MS: 394.3 (M+H).

Example 524-(4-(1H-tetrazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 4-(1H-tetrazol-1-yl)aniline instep 1. MS: 395.28 (M+H).

Example 532-((1R,2S)-2-aminocyclohexylamino)-4-(4-(oxazol-5-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 4-(oxazol-5-yl)aniline in step1.

Example 542-((1R,2S)-2-aminocyclohexylamino)-4-(3-(oxazol-4-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(oxazol-4-yl)aniline instep 1. MS: 394.28 (M+H).

Example 554-(3-(1H-1,2,4-triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine 5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-(1H-1,2,4-triazol-1-yl)aniline in step 1. MS: 394.2 (M+H).

Example 562-((1R,2S)-2-aminocyclohexylamino)-4-(3-(5-methyl-1,2,4-oxadiazol-3-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-(5-methyl-1,2,4-oxadiazol-3-yl)aniline in step 1. MS: 409.28 (M+H).

Example 572-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2-methylthiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(2-methylthiazol-4-yl)anilinein step 1. MS: 424.37 (M+H).

Example 582-((1R,2S)-2-aminocyclohexylamino)-4-(4-(5-methyl-1,2,4-oxadiazol-3-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using4-(5-methyl-1,2,4-oxadiazol-3-yl)aniline in step 1. MS: 409.5 (M+H).

Example 594-(3-(1H-pyrazol-5-yl)phenylamino)-2-(1R,2S)-2-aminocyclohexyl amino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(1H-pyrazol-5-yl)aniline instep 1. MS: 393.0 (M+H).

Example 602-((1R,2S)-2-aminocyclohexylamino)-4-(4-(thiophen-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 4-(thiophen-2-yl)aniline instep 1. MS: 410.0 (M+H).

Example 612-((1R,2S)-2-aminocyclohexylamino)-4-(3-methoxy-4-(oxazol-5-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-methoxy-4-(oxazol-5-yl)aniline in step 1. MS: 424.5 (M+H).

Example 622-((1R,2S)-2-aminocyclohexylamino)-4-(4-(3-methyl-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using4-(3-methyl-1H-pyrazol-1-yl)aniline in step 1. MS: 407.5 (M+H).

Example 632-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2-oxopyrrolidin-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using1-(3-aminophenyl)pyrrolidin-2-one in step 1. MS: 410.5 (M+H).

Example 642-((1R,2S)-2-aminocyclohexylamino)-4-(4-(3,5-dimethyl-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using4-(3,5-dimethyl-1H-pyrazol-1-yl)aniline in step 1. MS: 421.5 (M+H).

Example 652-((1R,2S)-2-aminocyclohexylamino)-4-(3-methoxy-5-(1H-tetrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-methoxy-5-(1H-tetrazol-1-yl)aniline in step 1. MS: 425.4 (M+H).

Example 664-(3-(1H-tetrazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(1H-tetrazol-1-yl)aniline instep 1. MS: 395.5 (M+H).

Example 672-((1R,2S)-2-aminocyclohexylamino)-4-(3-(5-methyl-1H-tetrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-(5-methyl-1H-tetrazol-1-yl)aniline in step 1. MS: 409.5 (M+H).

Example 682-((1R,2S)-2-aminocyclohexylamino)-4-(3-(1-methyl-1H-tetrazol-5-yl)phenylamino) pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using3-(1-methyl-1H-tetrazol-5-yl)aniline in step 1. MS: 409.5 (M+H).

Example 692-((1R,2S)-2-aminocyclohexylamino)-4-(4-(5-methyl-1,2,4-oxadiazol-3-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using4-(5-methyl-1,2,4-oxadiazol-3-yl)aniline in step 1. MS: 409.5 (M+H).

Example 704-(3-(1H-pyrrol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(1H-pyrrol-1-yl)aniline instep 1. MS: 392.5 (M+H).

Example 714-(4-(1H-pyrrol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 4-(1H-pyrrol-1-yl)aniline instep 1. MS: 392.4 (M+H).

Example 722-((1R,2S)-2-aminocyclohexylamino)-4-(3-(5-methylfuran-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(5-methylfuran-2-yl)anilinein step 1. MS: 407.5 (M+H).

Example 732-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyrrolidin-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-(pyrrolidin-1-yl)aniline instep 1. MS: 396.6 (M+H).

Example 742-((1R,2S)-2-aminocyclohexylamino)-4-(3-morpholinophenylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 3-morpholinoaniline in step 1.MS: 412.5 (M+H).

Example 754-(4-(1,3,4-oxadiazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of compound 122, and using 4-(1,3,4-oxadiazol-2-yl)anilinein step 1. MS: 395.3 (M+H).

Example 764-(3-(2H-tetrazol-5-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

Compound 147.4:2-((1R,2S)-2-aminocyclohexylamino)-4-(3-cyanophenylamino)pyrimidine-5-carboxamide was synthesized using the procedure describedfor the synthesis of Example 77. MS: 352.2 (M+H).

Compound 147:4-(3-(2H-tetrazol-5-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamidewas synthesized by reacting compound 147.4 (75 mg, 0.213 mmol), sodiumazide (90 mg, 1.384 mmol, 6.5 equiv), and zinc bromide (53 mg, 0.235mmol) in isopropanol/water (1:1) (4 mL) heated to reflux for 14 hrs,when an HPLC analysis showed a complete reaction. Then cooled thereaction mixture to 0° C., and acidified with 3N HCl to pH 1, filteredthrough celite, concentrated the filtrate to dryness to yield the titlecompound, 179, purified it by RP-HPLC. MS: 395.1 (M+H), 417.2 (M+Na).

Example 784-(4-(2H-tetrazol-5-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

This compound was synthesised using the synthetic scheme described forthe synthesis of Example 5, and using 4-cyanoaniline in step 1. MS:395.1 (M+H), 417.2 (M+Na).

Example 792-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(3-(trifluoromethyl)-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of trifluoromethylpyrazole 149.1 (1.00 g, 7.35 mmol),3,4-difluoro-1-nitrobenzene 149.2 (0.68 mL, 6.13 mmoL) and cesiumcarbonate (3.00 g, 9.2 mmol) in 20 mL dry DMF was stirred in 50° C. bathfor 4 hours. It was diluted with 300 mL ethyl acetate and washed withwater 4 times. The organic phase was dried over MgSO₄ and filtered toyield a solution of crude product 149.3. To this solution was addedcatalytic amount of 10%

Pd/C. To this stirred suspention was amounted a hydrogen balloon forovernight. The mixture was filtered and concentrated in vacuo to affordcrude aniline 149.4. MS found for C₁₀H₇F₄N₃ as (M+H)⁺ 246.3. It waspurified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with aniline 149.4 to replace aniline 74.1. MSfound for C₂₁H₂₂F₄N₈O as (M+H)⁺479.3. UV λ=243, 302 nm. NMR (CD₃OD): δ8.47 (s, 1H), 8.07 (broad s, 1H), 7.84 (m, 1H), 7.72 (m, 1H), 7.39 (m,1H), 6.74 (d, J=2.0 Hz, 1H), 4.33 (m, 1H), 3.66 (m, 1H), 1.83-1.49 (m,8H) ppm.

Example 802-((1R,2S)-2-aminocyclohexylamino)-4-(3,4-bis(3-(trifluoromethyl)-1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of trifluoromethylpyrazole 150.1 (2.00 g, 14.7 mmol),3,4-difluoro-1-nitrobenzene 150.2 (1.17 g, 7.3 mmoL) and cesiumcarbonate (5.5 g, 17 mmol) in 30 mL dry DMF was stirred in 50° C. bathfor overnight. It was diluted with 300 mL ethyl acetate and washed withwater four times. The organic phase was dried over MgSO₄ and filtered toyield a solution of crude product 150.3. To this solution was addedcatalytic amount of 10% Pd/C. To this stirred suspention was amounted ahydrogen balloon for overnight. The mixture was filtered andconcentrated in vacuo to afford crude aniline 150.4. MS found forC₁₄H₉F₆N₅ as (M+H)⁺ 362.2. It was purified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 150.4 to replace aniline-74.1. MSfound for C₂₅H₂₄F₆N₁₀O as (M+H)⁺595.3. UV λ=246, 301 nm. NMR (CD₃OD): δ8.61 (s, 1H), 8.45 (broad s, 1H), 7.78-7.65 (m, 4H), 6.75 (d, J=2.0 Hz,1H), 6.72 (d, J=2.4 Hz, 1H), 4.41 (m, 1H), 3.59 (m, 1H), 1.86-1.48 (m,8H) ppm.

Example 812-((1R,2S)-2-aminocyclohexylamino)-4-(3-morpholino-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of pyrazole 151.1 (0.50 g, 7.4 mmol),3,4-difluoro-1-nitrobenzene 151.2 (0.68 mL, 6.1 mmoL) and cesiumcarbonate (3.0 g, 9.2 mmol) in 15 mL dry NMP was stirred in a sealedtube in 80° C. bath for 3 hours to give compound 151.3. To the reactionsealed tube was then added morpholine (1.6 mL, 18.4 mmol). The mixturewas stirred in 120° C. bath for over 24 hours. It was diluted with 300mL ethyl acetate and washed with water four times. The organic phase wasdried over MgSO₄ and filtered to yield a solution of crude product151.4. To this solution was added catalytic amount of 10% Pd/C. To thisstirred suspention was amounted a hydrogen balloon for overnight. Themixture was filtered and concentrated in vacuo to afford crude aniline151.5. MS found for C₁₃H₁₆N₄O as (M+H)⁺245.2. It was purified usingflash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with aniline 151.5 to replace aniline 74.1. MSfound for C₂₄H₃₁N₉O₂ as (M+H)⁺ 478.3. UV λ=247 nm.

Example 824-(3-(1H-imidazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The mixture of imidazole 152.1 (0.64 g, 9.4 mmol),3-fluoro-1-nitrobenzene 152.2 (0.50 mL, 4.7 mmoL) and cesium carbonate(3.1 g, 9.4 mmol) in 15 mL dry NMP was stirred in a sealed tube in 120°C. bath for 3 hours. It was diluted with 300 mL ethyl acetate and washedwith water four times. The organic phase was dried over MgSO₄ andfiltered to yield a solution of crude product 152.3. To this solutionwas added catalytic amount of 10% Pd/C. To this stirred suspention wasamounted a hydrogen balloon for overnight. The mixture was filtered andconcentrated in vacuo to afford crude aniline 152.4. MS found for C₉H₉N₃as (M+H)⁺ 160.1.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 152.4 to replace aniline 74.1. MSfound for C₂₄H₃₁N₉O₂ as (M+H)⁺ 478.3. UV λ=241 nm.

Example 832-((1R,2S)-2-aminocyclohexylamino)-4-(4-(5-methyl-1,3,4-thiadiazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: Iodobenzene 77.2 (500 mg, 1.2 mmol) was dissolved in 12 mLdioxane in a sealed tube. To it were added bromothiadiazole 153.1 (240mg, 1.3 mmol), hexamethylditin (0.25 mL, 1.2 mmol) and Pd(PPh₃)₄ (280mg, 0.24 mmol). The mixture was degassed using argon stream for 3minutes and stirred in 110° C. bath for 90 min. It was concentrated invacuo and subjected to silica flash column to isolate compound 153.2(130 mg, 28%). MS found for C₁₇H₁₇N₅O₂S₂ as (M+H)⁺ 388.1.

Step 2: Ethyl ester 153.2 (130 mg, 0.34 mmol) was dissolved in 20 mLTHF. To it were added lithium hydroxide hydrate (42 mg, 1.0 mmol) and 3mL water. The mixture was stirred for 2 hours at RT. It was concentratedin vacuo to remove THF and carefully treated with 1N HCl till pHreaching 3. A yellow solid crashed out from the solution. It wasisolated using a Büchner funnel, washed with cold water, dried in vacuumoven to give compound 153.3. MS found for C₁₅H₁₃N₅O₂S₂ as (M+H)⁺ 360.1.

Step 3: Carboxylic acid 153.3 (0.34 mmol) was dissolved in 10 mL DMF. Toit were added EDC hydrochloride (100 mg, 0.51 mmol) and HOBt hydrate (70mg, 0.51 mmol). The mixture was stirred at RT for 1 hour. To it was thenadded ammonia (commercial 0.5N solution in dioxane, 3.4 mL, 1.7 mmol).The mixture was stirred for overnight. It was then concentrated in vacuoto remove dioxane. The mixture was then subjected to reverse phasepreparative HPLC to isolate compound 153.4 (22 mg, 18% for 2 steps). MSfound for C₁₅H₁₄N₆OS₂ as (M+H)⁺ 359.1.

Step 4: Compound 153.4 (22 mg, 0.06 mmol) was dissolved in 5 mL NMP. Toit was added MCPBA (65% pure, 21 mg, 0.072 mmol). It was stirred at RTfor 2 hours. To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate 81.2 (0.3 M, 0.4 mL, 0.12 mmol) andDIEA (31 μL, 0.18 mmol). The mixture was stirred for 80 minutes at 90°C. bath. This mixture was diluted with ethyl acetate, washed withsaturated Na₂CO₃ aqueous solution twice and water. The organic phase wasdried over MgSO₄ and concentrated in vacuo to afford crude compound153.5. MS found for C₂₅H₃₂N₈O₃S as (M+H)⁺ 525.3.

Step 5: Compound 153.5 was stirred in a 1:1 mixture of TFA anddichloromethane at RT for 1 hour. It was concentrated in vacuo andsubjected to reverse phase preparative HPLC to isolate the titlecompound. MS found for C₂₀H₂₄N₈OS as (M+H)⁺ 425.3. UV λ=241, 319 nm.

Example 842-((1R,2S)-2-aminocyclohexylamino)-4-(4-(2-methyl-1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of 2-methylimidazole 154.1 (1.52 g, 18.6 mmol),4-fluoro-1-nitrobenzene 154.2 (1.0 mL, 9.4 mmoL) and potassium carbonate(1.30 g, 9.4 mmol) in 20 mL dry DMF was stirred in 100° C. bath forovernight. It was diluted with 300 mL ethyl acetate and washed withwater 4 times. The organic phase was dried over MgSO₄ and filtered toyield a solution of crude product 154.3. To this solution was addedcatalytic amount of 10% Pd/C. To this stirred suspention was amounted ahydrogen balloon for overnight. The mixture was filtered andconcentrated in vacuo to afford crude aniline 154.4. MS found forC₁₀H₁₁N₃ as (M+H)⁺ 174.1. It was purified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 154.4 to replace aniline 74.1. MSfound for C₂₁H₂₆N₈O as (M+H)⁺407.3. UV λ=246, 293 nm. NMR (CD₃OD): δ8.60 (s, 1H), 7.96 (m, 2H), 7.70 (d, J=8.4 Hz, 2H), 7.66-7.61 (m, 4H),4.47 (m, 1H), 3.73 (m, 1H), 2.60 (s, 3H), 1.92-1.59 (m, 8H) ppm.

Example 852-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2-methyl-1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of 2-methylimidazole 155.1 (0.77 g, 9.4 mmol),3-fluoro-1-nitrobenzene 155.2 (0.50 mL, 4.7 mmoL) and cesium carbonate(3.07 g, 9.4 mmol) in 15 mL dry NMP was stirred in 120° C. bath for 6hours. It was diluted with 300 mL ethyl acetate and washed with water 4times. The organic phase was dried over MgSO₄ and filtered to yield asolution of crude product 155.3. To this solution was added catalyticamount of 10% Pd/C. To this stirred suspention was amounted a hydrogenballoon for overnight. The mixture was filtered and concentrated invacuo to afford crude aniline 155.4. MS found for C₁₀H₁₁N₃ as (M+H)⁺174.1. It was purified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 155.4 to replace aniline 74.1. MSfound for C₂₁H₂₆N₈O as (M+H)⁺ 407.3. UV λ=243, 285 nm. NMR (CD₃OD): δ8.58 (s, 1H), 8.06 (m, 1H), 7.83 (m, 1H), 7.73-7.71 (m, 2H), 7.64 (m,1H), 7.44 (m, 1H), 4.41 (m, 1H), 3.66 (m, 1H), 2.62 (s, 3H), 1.89-1.55(m, 8H) ppm.

Example 864-(3-(1H-1,2,3-triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The mixture of 1H-1,2,3-triazole 156.1 (0.55 mL 9.4 mmol),3-fluoro-1-nitrobenzene 156.2 (0.50 mL, 4.7 mmoL) and cesium carbonate(3.07 g, 9.4 mmol) in 15 mL dry NMP was stirred in 120° C. bath for 17hours. It was diluted with 300 mL ethyl acetate and washed with water 4times. The organic phase was dried over MgSO₄ and filtered to yield asolution of crude products 156.3 and 156.4 in nearly 1:1 ratio. To thissolution was added catalytic amount of 10% Pd/C. To this stirredsuspention was amounted a hydrogen balloon for overnight. The mixturewas filtered and concentrated in vacuo to afford crude anilines 156.5and 156.6. MS found for C₈H₈N₄ as (M+H)⁺ 161.1. The two anilines werepurified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 156.5 to replace aniline 74.1. MSfound for C₁₉H₂₃N₉O as (M+H)⁺ 394.3. UV λ=244 nm. NMR (CD₃OD): δ 8.90(s, 1H), 8.63 (d, J=1.2 Hz, 1H), 8.57 (s, 1H), 7.95 (d, J=1.2 Hz, 1H),7.62-7.58 (m, 2H), 7.43 (m, 1H), 4.68 (m, 1H), 3.73 (m, 1H), 1.91-1.53(m, 8H) ppm.

Example 874-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 156.6 (shown in Example 86) toreplace aniline 74.1. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.3. UV λ=250nm. NMR (CD₃OD): δ 8.77 (s, 1H), 8.46 (s, 1H), 7.88 (s, 2H), 7.83 (d,J=7.6 Hz, 1H), 7.45 (dd, J=8.4, 8.0 Hz, 1H), 7.23 (d, J=8.0 Hz, 1H),4.52 (m, 1H), 3.58 (m, 1H), 1.82-1.43 (m, 8H) ppm.

Example 882-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-(hydroxymethyl)-1H-1,2,3-triazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in the scheme (Example 45) with propargyl alcohol toreplace trimethylsilylacetylene. MS found for C₂₀H₂₅N₉O₂ as (M+H)⁺424.3. UV λ=242, 300 nm.

Example 892-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-(aminomethyl)-1H-1,2,3-triazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in the Scheme (Example 45) with propargylamine to replacetrimethylsilylacetylene. MS found for C₂₀H₂₆N₁₀O as (M+H)⁺ 423.3. UVλ=242, 301 nm.

Example 902-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-carbamoyl-1H-1,2,3-triazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in the scheme (Example 45) with propynoic acid amide toreplace trimethylsilylacetylene. MS found for C₂₀H₂₄N₁₀O₂ as (M+H)⁺437.3. UV λ=242, 300 nm.

Example 912-((1R,2S)-2-aminocyclohexylamino)-4-(4-(methylsulfonyl)-3-morpholinophenylamino)pyrimidine-5-carboxamide

The mixture of sodium thiomethoxide (2.85 g, 40.6 mmol) and3,4-difluoro-1-nitrobenzene 161.1 (3.0 mL, 27.1 mmoL) in 20 mL dry NMPwas stirred at RT for 3 hours. It was diluted with 300 mL ethyl acetateand washed with water four times. The organic phase was dried overMgSO₄, concentrated in vacuo and dissolved in 270 mL DCM. To it wasadded MCPBA (65%, 14.3 g, 54 mmol) in small portions. The mixture wasstirred for 2 hours at RT, diluted with more DCM, washed with 0.1 N NaOHthree times and brine. This solution was dried over MgSO₄ andconcentrated in vacuo to give crude 161.2.

Crude 161.2 (490 mg, 2.2 mmol) was dissolved in 10 mL dry NMP. To it wasadded morpholine (1.2 mL, 6.6 mmol). The mixture was stirred at 60° C.bath for 1 hour. It was diluted by ethyl acetate 300 mL, washed withbrine three times, dried over MgSO₄ and filtered to yield a solution ofcrude product 161.3. To this solution was added catalytic amount of 10%Pd/C. To this stirred suspention was amounted a hydrogen balloon forovernight. The mixture was filtered and concentrated in vacuo to affordcrude aniline 161.4. MS found for C₁₁H₁₆N₂O₃S as (M+H)⁺ 257.1. It waspurified using flash column.

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with aniline 161.4 to replace aniline 74.1. MSfound for C₂₂H₃₁N₇O₄S as (M+H)⁺ 490.3. UV λ=249, 301 nm.

Example 922-((1R,2S)-2-aminocyclohexylamino)-4-(4-fluoro-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: The mixture of 3-bromo-4-fluoroaniline (860 mg, 4.53 mmol),1,2,3-triazole (1.05 mL, 18.1 mmol), K₃PO₄ (1.92 g, 9.06 mmol), CuI (430mg, 2.27 mmol), N,N′-dimethylethylenediamine (0.29 mL, 2.72 mmol) in 10mL dioxane and 5 mL DMSO was stirred in a sealed tube at 120° C. for 5days. A mixture of 243.1 and 243.2 (in 1.5:1 ratio) along with leftoverstarting materials were obtained. To the mixture was poured EtOAc 250mL. It was vigoriously stirred, washed with water and brine, dried overMgSO₄, filtered, concentrated in vacuo and subjected to flash column.Compound 243.1 left the column at 70% EtOAc in hexane and compound 243.2at 90% EtOAc in hexane. Step 2: To the mixture of aniline 243.1 (255 mg,1.44 mmol) and ethyl 4-chloro-2-methylthio-5-pyrimidinecarboxylate(243.3, CAS 5909-24-0, 336 mg, 1.44 mmol) in 15 mL DMF was added DIEA(0.5 mL, 2.88 mmol). The mixture was stirred at 85° C. for 5 hours. Toit was added 250 mL EtOAc, washed with brine x3, dried, filtered andconcentrated in vacuo to afford compound 243.4 in quantatititive yield.MS found for C₁₆H₁₅FN₆O₂S as (M+H)⁺ 375.1.

Step 3: The above prepapred compound (1.44 mmol) was dissolved in 80 mLTHF and 10 mL water. To it was added LiOH hydrate (302 mg, 7.2 mmol).The mixture was stirred for overnight. It was concentrated in vacuo toremove THF. To the residue was added 1N HCl till pH reaching 2. Thesolid product was isolated by filtration. It was washed thoroughly withcold water and dried in vacuum oven to give compound 243.5 inquantatitive yield as a tan solid. MS found for C₁₄H₁₁FN₆O₂S as (M+H)⁺347.1.

Step 4: The above prepared compound 243.5 (468 mg, 1.35 mmol) wasstirred in 25 mL DMF. To it were added EDC.HCl (390 mg, 2.03 mmol) andHOBt hydrate (311 mg, 2.03 mmol). The mixture was stirred for 1 hr andHPLC indicated that all the starting 243.5 had been consumed. To thismixture was then added ammonium (0.5N solution in dixoane, 8.1 mL, 4.05mmol). The mixture was stirred for 3 hrs and concentrated in vacuo toremove dioxane. Water was added to the residue and solid crashed out.This solid was isolated by filtration, washed with cold water thoroughlyand dried in vacuum oven to give compound 243.6 as a tan solid. MS foundfor C₁₄H₁₂FN₇OS as (M+H)⁺ 346.1.

Step 5: The above prepared compound 243.6 (100 mg, 0.29 mmol) wasdissolved in 5 mL NMP. To it was added MCPBA (65% pure, 92 mg, 0.35mmol). It was stirred at RT for 30 minutes to afford a mixture of thecorresponding sulfoxide and sulfone. To it then were added DIEA (160 μL,0.90 mmol) and then tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (81.2,94 mg, 0.45 mmol). The mixture was stirred for 3 hrs at 90° C. bath. Itwas then diluted with 150 mL EtOAc, washed with sat Na₂CO₃ and brine x2,dried over MgSO₄, filtered, and concentrated in vacuo. The residue wasthen exposed to 1:1 mixture of TFA and DCM for 1 hr. It was concentratedand subjected to reverse phase HPLC to isolate the title compound 243.MS found for C₁₉H₂₂FN₉O as (M+H)⁺ 412.3. UV λ=247 nm. NMR (CD₃OD): δ8.58 (m, 1H), 8.55 (s, 1H), 8.05 (s, 2H), 7.45-7.43 (m, 2H), 4.53 (m,1H), 3.64 (m, 1H), 1.86-1.54 (m, 8H) ppm.

Example 932-((1R,2S)-2-aminocyclohexylamino)-4-(4-cyano-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: 2-fluoro-4-nitrobenzonitrile (2.98 g, 18 mmol) was dissolved in40 mL NMP. To it were added 1,2,3-triazole (1.1 mL, 18.8 mmol) and K₂CO₃(2.98 g, 21.6 mmol). The mixture was stirred at RT for 20 h to givecompound 230.1 and 230.2 (less polar) in 1.3:1 ratio. The mixture wastaken into ethyl acetate, washed with water and brine x3. It was dried,concentrated and subjected to silica flash column to isolate desiredcompound 230.2 using 20% ethyl acetate in hexane.

Step 2: Compound 230.2 from Step 1 was dissolved in 300 mL ethylacetate. To it was added 500 mg 10% Pd/C. The mixture was placed on Parrshaker under 45 psi hydrogen pressure for overnight. The mixture wasfiltered through celite, which was thoroughly washed using methanol. Thefiltrate was concentrated and subjected to silica flash column toisolate aniline 230.3 (0.59 g, 18% for 2 steps).

Step 3: Compound 230.4 (0.66 g, 3.0 mmol) was dissolved in 20 mL NMP. Toit were added compound 230.3 (0.55 g, 3.0 mmol) and then DIEA (0.78 mL,4.5 mmol). The mixture was stirred at 75° C. for 24 h. To it was thenadded NaSMe (0.42 g, 6.0 mmol). The mixture was stirred at 70° C. forovernight. The mixture was diluted with ethyl acetate, washed withbrine×4, dried, concentrated in vauo. The residue was then dissolved in50 mL THF. To it were added LiOH.H₂O (1.26 g, 30 mmol) and 50 mL water.The mixture was stirred for 90 min at RT. It was acidified using 6 N HCltill pH reaching 2. The solid precipitate was collected by filtration.It was washed with water and dried in vacuo oven. It was crude compound230.5.

The title compound was made using the similar chemistry scheme shown forExample 93 using compound 230.5. MS found for C₂₀H₂₂N₁₀O as (M+H)⁺419.4. UV λ=259, 314 nm.

Example 942-((1R,2S)-2-aminocyclohexylamino)-4-(3-cyano-5-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 92. MS found for C₂₀H₂₂N₁₀O as (M+H)⁺ 419.3. UV λ=252 nm. NMR(CD₃OD): δ 9.03 (s, 1H), 8.61 (s, 1H), 8.16 (m, 1H), 8.02 (s, 2H), 7.91(m, 1H), 4.62 (m, 1H), 3.69 (m, 1H), 1.94-1.58 (m, 8H) ppm.

Example 952-((1R,2S)-2-aminocyclohexylamino)-4-(3-cyano-5-(1H-1,2,3-triazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 93. MS found for C₂₀H₂₂N₁₀O as (M+H)⁺ 419.4. UV λ=244, 288 nm.NMR (CD₃OD): δ 9.15 (s, 1H), 8.71 (s, 1H), 8.63 (s, 1H), 7.99-7.97 (m,3H), 4.67 (m, 1H), 3.72 (m, 1H), 1.90-1.58 (m, 8H) ppm.

Example 962-((1R,2S)-2-aminocyclohexylamino)-4-(3-(6-methoxypyridin-2-yl)phenylamino)pyrimidine-5-carboxamide

Commercially available compound 233.1 (2.3 g, 12.2 mmol) and boronicacid 233.2 (1.68 g, 12.2 mmol) were placed in 40 mL dioxane and 20 mLwater. To it were added K₂CO₃ (5.05 g, 36.6 mmol) and Pd(Ph₃P)₂Cl₂ (0.86g, 1.22 mmol). The mixture was bubbled with argon stream for 3 min andsent to 85° C. bath under argon. The mixture was stirred for 90 min. Itwas concentrated in vacuo to remove dioxane. The mixture then wasextracted with chloroform x4. The organic extract was dried andconcentrated in vauo to afford crude aniline 233.3.

The title compound was then prepared using the similar chemistry schemeshown for Example 86 using aniline 233.3. MS found for C₂₃H₂₇N₇O₂ as(M+H)⁺ 434.4. UV λ=245, 296 nm.

Example 972-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methoxypyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the similar chemistry scheme shownfor Example 96. MS found for C₂₂H₂₆N₈O₂ as (M+H)⁺ 435.4. UV λ=250 nm.NMR (CD₃OD): 8.53 (s, 1H), 8.42 (s, 1H), 7.92 (m, 1H), 7.77 (m, 1H),7.54-7.50 (m, 3H), 6.79 (d, J=8.0 Hz, 1H), 4.30 (m, 1H), 4.01 (s, 3H),3.64 (m, 1H), 1.85-1.36 (m, 8H) ppm.

Example 982-((1R,2S)-2-aminocyclohexylamino)-4-(3-(5-fluoropyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the similar chemistry scheme shownfor Example 96. MS found for C₂₁H₂₃FN₈O as (M+H)⁺ 423.3. UV λ=249

Example 992-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methyl-1H-1,2,3-triazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 86. MS found for C₂₀H₂₅N₉O as (M+H)⁺ 408.4. UVλ=245 nm.

Example 1002-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methyl-2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 87. MS found for C₂₀H₂₅N₉O as (M+H)⁺ 408.4. UVλ=251 nm.

Example 101a4-(3-(2H-tetrazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The mixture of 3-iodoaniline (0.5 mL, 4.2 mmol), tetrazole (0.88 g, 12.6mmol), K₃PO₄ (2.67 g, 12.6 mmol), CuI (400 mg, 2.1 mmol), DMEDA (0.27mL, 2.5 mmol) in 6 mL dioxane and 6 mL DMSO was stirred at 105° C. in asealed flask for 3 days to cleanly give a mixture of aniline 238.1 (lesspolar) and 238.2 in ratio of 17:1 (determined by HPLC). It was dilutedwith ethyl acetate, washed with water and brine x2. It was dried,concentrated and subjected to silica flash column to isolate aniline238.1 using 40% ethyl acetate in hexane.

The title compound was prepared using the same synthetic schemedemonstrated in Example 86. MS found for C₁₈H₂₂N₁₀O as (M+H)⁺ 395.4. UVλ=250 nm. NMR (CD₃CN): δ 9.02 (s, 1H), 8.87 (s, 1H), 7.98 (d, J=8.0 Hz,1H), 7.67-7.53 (m, 3H), 4.83 (m, 1H), 3.67 (m, 1H), 1.8-1.4 (m, 8H) ppm.

Example 101b2-((1R,2S)-2-aminocyclohexylamino)-4-(3-methyl-5-(2H-tetrazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of 3-bromo-5-methylaniline. HCl (0.94 g, 4.2 mmol),tetrazole (0.88 g, 12.6 mmol), K₃PO₄ (4.45 g, 21 mmol), CuI (400 mg, 2.1mmol), DMEDA (0.27 mL, 2.5 mmol) in 6 mL dioxane and 6 mL DMSO wasstirred at 105° C. in a sealed flask for 3 days to cleanly giveexclusively aniline 239.1. Aniline 239.2 was not found by HPLC/LCMS. Itwas diluted with ethyl acetate, washed with water and brine x2. It wasdried, concentrated and subjected to silica flash column to isolateaniline 239.1 using 40% ethyl acetate in hexane.

The title compound was prepared using the same synthetic schemedemonstrated in Example 86. MS found for C₁₉H₂₄N₁₀O as (M+H)⁺ 409.4. UVλ=250 nm. NMR (CD₃CN): δ 8.85 (s, 1H), 8.80 (s, 1H), 8.58 (s, 1H), 7.83(s, 1H), 4.83 (m, 1H), 3.68 (m, 1H), 2.50 (s, 3H), 1.8-1.4 (m, 8H) ppm.

Example 1022-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-methyl-4H-1,2,4-triazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 86 with the corresponding aniline. MS found forC₂₀H₂₅N₉O as (M+H)⁺ 408.4. UV λ=246, 304 nm.

Example 1034-(3-(1H-benzo[d]imidazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The mixture of 3-iodoaniline (0.5 mL, 4.2 mmol), benzimidazole (1.5 g,12.6 mmol), K₃PO₄ (2.67 g, 12.6 mmol), CuI (400 mg, 2.1 mmol), DMEDA(0.27 mL, 2.5 mmol) in 6 mL dioxane and 6 mL DMSO was stirred in asealed flask at 120° C. for 2 days to cleanly afford aniline 241.1. Themixture was diluted with 300 mL ethyl acetate, washed with water andbrine, dried, filtered, concentrated, subjected to silica flash columnwith 70% ethyl acetate in hexane to isolate aniline 241.1 (730 mg, 83%)as white solid.

The title compound was prepared using the same synthetic schemedemonstrated in Example 89 with the aniline. MS found for C₂₄H₂₆N₈O as(M+H)⁺ 443.4. UV λ=249 nm. NMR (CD₃OD): δ 9.07 (s, 1H), 8.56 (s, 1H),8.29 (s, 1H), 7.87 (m, 1H), 7.74 (m, 3H), 7.56 (m, 3H), 4.14 (m, 1H),3.50 (m, 1H), 1.72-1.11 (m, 8H) ppm.

Example 1044-(3-(1H-indazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide(242.A) and4-(3-(2H-indazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide(242.B)

The mixture of 3-iodoaniline (0.5 mL, 4.2 mmol), indazole (1.5 g, 12.6mmol), K₃PO₄ (2.67 g, 12.6 mmol), CuI (400 mg, 2.1 mmol), DMEDA (0.27mL, 2.5 mmol) in 6 mL dioxane and 6 mL DMSO was stirred in a sealedflask at 120° C. for 17 h to cleanly afford aniline 242.1 (less polar)and aniline 242.2 in ratio of 6.9:1 (determined by HPLC). The mixturewas diluted with 300 mL ethyl acetate, washed with water and brine,dried, filtered, concentrated, subjected to silica flash column toisolate the two anilines.

The two title compounds were prepared using the same synthetic schemedemonstrated in Example 86 with the corresponding anilines. With aniline242.1, compound 242.A was prepared: MS found for C₂₄H₂₆N₈O as (M+H)⁺443.4. UV λ=247, 301 nm. With aniline 242.2, compound 242.B wasprepared: MS found for C₂₄H₂₆N₈O as (M+H)⁺ 443.4. UV λ=240, 295 nm.Compound 104A: NMR (CD₃OD): δ 8.57 (m, 1H), 8.56 (s, 1H), 8.31 (s, 1H),7.87 (m, 1H), 7.83 (m, 3H), 7.59 (m, 2H), 7.51 (m, 1H), 7.33 (m, 1H),7.28 (m, 1H), 4.34 (m, 1H), 3.60 (m, 1H), 1.80-1.22 (m, 8H) ppm.Compound 104B: NMR (CD₃OD): δ 8.84 (s, 1H), 8.78 (m, 1H), 8.55 (s, 1H),7.78-7.76 (m, 2H), 7.68 (d, J=9.2 Hz, 1H), 7.57 (m, 1H), 7.42-7.34 (m,2H), 7.13 (m, 1H), 4.46 (m, 1H), 3.63 (m, 1H), 1.85-1.19 (m, 8H) ppm.

Example 1054-(3-(1H-benzo[d][1,2,3]triazol-1-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide(243.A) and4-(3-(2H-benzo[d][1,2,3]triazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide(243.B)

The mixture of 3-iodoaniline (0.5 mL, 4.2 mmol), indazole (1.5 g, 12.6mmol), K₃PO₄ (2.67 g, 12.6 mmol), CuI (400 mg, 2.1 mmol), DMEDA (0.27mL, 2.5 mmol) in 6 mL dioxane and 6 mL DMSO was stirred in a sealedflask at 120° C. for 17 h to cleanly afford aniline 243.1 and aniline243.2 (less polar) in ratio of 7.5:1 (determined by HPLC). The mixturewas diluted with 300 mL ethyl acetate, washed with water and brine,dried, filtered, concentrated, subjected to silica flash column toisolate the two anilines.

The two title compounds were prepared using the same synthetic schemedemonstrated in Example 86 with the corresponding anilines. With aniline243.1, compound 2432.A was prepared: MS found for C₂₃H₂₅N₉O as (M+H)⁺444.4. UV λ=246, 291 nm. With aniline 243.2, compound 243.B wasprepared: MS found for C₂₃H₂₅N₉O as (M+H)⁺ 444.4. UV λ=234, 303 nm.Compound 105A: NMR (CD₃OD): δ 8.78 (m, 1H), 8.56 (s, 1H), 8.12 (d, J=8.4Hz, 1H), 7.95 (d, J=8.4 Hz, 1H), 7.68 (m, 3H), 7.54 (m, 1H), 7.47 (m,1H), 4.57 (m, 1H), 3.68 (m, 1H), 1.90-1.44 (m, 8H) ppm. Compound 105B:NMR (CD₃OD): δ 9.07 (s, 1H), 8.55 (s, 1H), 8.16 (d, J=7.6 Hz, 1H), 7.92(m, 2H), 7.59 (m, 1H), 7.49 (m, 2H), 7.42 (m, 1H), 4.61 (m, 1H), 3.67(m, 1H), 1.93-1.59 (m, 8H) ppm.

Example 1062-((1R,2S)-2-aminocyclohexylamino)-4-(3-(imidazo[1,2-a]pyridin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 86 with the biaryl aniline (commerciallyavailable). MS found for C₂₄H₂₆N₈O as (M+H)⁺ 443.4. UV λ=240, 292 nm.

Example 1074-(3-(2H-benzo[b][1,4]oxazin-4(3H)-yl)phenylamino)-2-(1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The mixture of commercially available compound 245.1 (600 mg, 4.44mmol), 3-iodo-1-nitrobenzene (245.2, 1.11 g, 4.44 mmol), Pd(dba)₂ (128mg, 0.22 mmol), Ph₅FcP(tBu)₂ (313 mg, 0.44 mmol) and NaOtBu (640 mg,0.66 mmol) in 8 mL toluene was stirred at 50° C. under argon forovernight (21 h). Compound 245.2 was formed cleanly. The mixture wasdiluted with 300 mL ethyl acetate, washed with brine x3, dried, filteredthrough a thin silica plug. The filtrate was subjected to treatment of200 mg 10% Pd/C at 50 psi H₂ Parr shaker for overnight. The mixture wasfiltered through celite, and the celite was thoroughly washed withmethanol. The filtrate was concentrated in vacuo to afford crude aniline245.4 (870 mg, 87% for 2 steps).

The title compound was prepared using the same synthetic schemedemonstrated in Example 12 with the aniline prepared above. MS found forC₂₅H₂₉N₇O₂ as (M+H)⁺ 460.4. UV λ=243, 290 nm.

Example 1082-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-phenylpiperazin-1-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of 3-fluoro-1-nitrobenzene (1.0 mL, 9.4 mmol),N-phenylpiperazine (2.87 mL, 18.8 mmol) and cesium carbonate (6.13 g,18.8 mmol) in NMP was stirred at 128° C. for three nights. It wasdiluted with ethyl acetate, washed with brine x2, dried, concentratedand purified using silica flash column to afford compound 245.1 (1.19 g,45%). It was dissolved in 200 mL ethyl acetate and treated with 500 mg10% Pd/C under 50 psi hydrogen pressure on Parr shaker for overnight.The mixture was filtered through celite, and the celite was washedthoroughly with methanol. The filtrate was concentrated in vacuo toafford aniline 245.2.

The title compound was prepared using the same synthetic schemedemonstrated in Example 12 with the aniline prepared above. MS found forC₂₇H₃₄N₈O as (M+H)⁺ 487.5. UV λ=245 nm.

Example 1092-((1R,2S)-2-aminocyclohexylamino)-4-(3-(quinolin-6-yl)phenylamino)pyrimidine-5-carboxamide

The mixture of 6-bromoquinoline (870 mg, 4.2 mmol), boronic acid (580mg, 4.2 mmol), Pd(Ph₃P)₂Cl₂ (590 mg, 0.84 mmol), K₂CO₃ (1.74 g, 12.6mmol) in 20 mL dioxane and 10 mL water was degassed using argon streamfor 3 min and stirred at 85° C. under argon for 90 min. The mixture wasdiluted with ethyl acetate, washed with water and brine, dried,concentrated, subjected to silica flash column with 60% ethyl acetate inhexane to isolate the featured aniline.

The title compound was prepared using the same synthetic schemedemonstrated in Example 86 with the aniline prepared above. MS found forC₂₆H₂₇N₇O as (M+H)⁺ 454.4. UV λ=263 nm.

Example 1102-((1R,2S)-2-aminocyclohexylamino)-4-(3-(quinolin-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 109. MS found for C₂₆H₂₇N₇O as (M+H)⁺ 454.4. UVλ=249 nm.

Example 1122-((1R,2S)-2-aminocyclohexylamino)-4-(4-(1-methyl-1H-pyrazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 83 with 3-iodo-1-methylpyrazole to replace2-bromo-5-methyl-1,3,4-thiadiazole 153.1. MS found for C₂₁H₂₆N₈O as(M+H)⁺ 407.3. UV λ=240, 311 nm.

Example 1132-((1R,2S)-2-aminocyclohexylamino)-4-(4-(1-methyl-1H-pyrazol-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 83 with 4-iodo-1-methylpyrazole to replace2-bromo-5-methyl-1,3,4-thiadiazole 153.1. MS found for C₂₁H₂₆N₈₀ as(M+H)⁺ 407.3. UV λ=239, 314 nm.

Example 1144-(4-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 87 with 4-fluoro-1-nitrobenzene to replace3-fluoro-1-nitrobenzene. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.3. UVλ=239, 310 nm. NMR (CD₃OD): δ 8.54 (s, 1H), 8.12 (m, 2H), 7.93 (s, 2H),7.81 (m, 2H), 4.40 (m, 1H), 3.73 (m, 1H), 1.94-1.58 (m, 8H) ppm.

Example 1152-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-5-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using 3-fluoro-5-iodoaniline using aprocedure same as that described in Example 92. MS found for C₁₉H₂₂FN₉Oas (M+H)⁺ 412.3. UV λ=250 nm.

Example 1162-((1R,2S)-2-aminocyclohexylamino)-4-(4-methoxy-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using 3-iodo-4-methoxyaniline using aprocedure same as that described in Example 92. 3-Iodo-4-methoxyanilinewas prepared from commercial 3-iodo-4-methoxy-1-nitrobenzene byhydrogenation with 5% sulfided Pt on carbon in EtOAc. MS found forC₂₀H₂₅N₉O₂ as (M+H)⁺ 424.3. UV λ=244, 295 nm.

Example 1172-((1R,2S)-2-aminocyclohexylamino)-4-(3-methoxy-5-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using 3-bromo-5-methoxyaniline using aprocedure same as that described in Example 92. 3-Bromo-5-methoxyanilinewas prepared from commercial 3-bromo-5-methoxy-1-nitrobenzene byhydrogenation with 5% sulfided Pt on carbon in EtOAc. MS found forC₂₀H₂₅N₉O₂ as (M+H)⁺ 424.3. UV λ=249 nm.

Example 1182-((1R,2S)-2-aminocyclohexylamino)-4-(4-methyl-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using4-methyl-3-(2H-1,2,3-triazol-2-yl)aniline using a procedure same as thatdescribed in Example 87. 4-Methyl-3-(2H-1,2,3-triazol-2-yl)aniline wasprepared from commercial 3-fluoro-4-methyl-1-nitrobenzene and1,2,3-triazole followed by hydrogenation with 10% Pd on carbon in EtOAc,as shown in Example 106. MS found for C₂₀H₂₅N₉O as (M+H)⁺ 408.3. UVλ=243, 281 nm. δ 1.5-1.9 (m, 8H), 2.36 (s, 3H), 3.60-3.70 (m, 1H),4.40-4.50 (m, 1H), 7.35-7.43 (m, 2H), 8.00 (s, 2H), 8.28-8.33 (m, 1H),8.53 (s, 1H)

Example 1192-((1R,2S)-2-aminocyclohexylamino)-4-(3-methyl-5-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial 3-bromo-5-methylanilineusing a procedure same as that described in Example 92. MS found forC₂₀H₂₅N₉O as (M+H)⁺ 408.3. UV λ=248 nm. δ 1.5-1.9 (m, 8H), 2.45 (s, 3H),3.65-3.75 (m, 1H), 4.6-4.7 (s, 1H), 7.12-7.18 (m, 1H), 7.72 (s, 1H),7.98 (s, 2H), 8.56 (s, 1H), 8.72-8.78 (m, 1H)

Example 1202-((1R,2S)-2-aminocyclohexylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

Step 1: The mixture of3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline (670 mg, 3.06mmol), 2-bromopyrimidine (486 mg, 3.06 mmol), Pd(PPh₃)₂Cl₂ (430 mg, 0.61mmol), K₂CO₃ (1.27 g, 9.18 mmol) in 20 mL dixoane and 10 mL water wasdegassed using Ar stream for 5 minutes and stirred at 85° C. under Arfor 2 hrs. It was diluted with 300 mL EtOAc and washed with water andbrine x2. The organic phase was then dried, concentrated in vacuo andsubjected to flash column to give compound 249.1 (white solid, 440 mg,84% yield, out of column with 60% EtOAc in hexane). MS found for C₁₀H₉N₃as (M+H)⁺ 172.1.

Step 2: To the mixture of aniline 249.1 (440 mg, 2.57 mmol) and ethyl4-chloro-2-methylthio-5-pyrimidinecarboxylate (243.3, CAS 5909-24-0, 600mg, 2.57 mmol) in 20 mL DMF was added DIEA (0.90 mL, 5.14 mmol). Themixture was stirred at 80° C. for 10 hours. To it was added 350 mLEtOAc, washed with brine x2, dried, filtered and concentrated in vacuoto afford compound 249.2 in quantatititive yield. MS found forC₁₈H₁₇N₅O₂S as (M+H)⁺ 368.1.

Step 3: The above prepapred compound (2.57 mmol) was dissolved in 50 mLTHF and 5 mL water. To it was added LiOH hydrate (540 mg, 12.9 mmol).The mixture was stirred for overnight. It was concentrated in vacuo toremove THF. To the residue was added 1N HCl until pH reaching 2. Thesolid product was isolated by filtration. It was washed thoroughly withcold water and dried in vacuum oven to give compound 249.3 inquantatitive yield as a tan solid. MS found for C₁₆H₁₃N₅O₂S as (M+H)⁺340.1.

Step 4: The above prepared compound 249.3 (2.57 mmol) was stirred in 20mL DMF. To it were added EDC.HCl (740 mg, 3.86 mmol) and HOBt hydrate(590 mg, 3.86 mmol). The mixture was stirred for 1 hr and HPLC indicatedthat all the starting 249.3 had been consumed. To this mixture was thenadded ammonium (0.5N solution in dixoane, 15 mL, 7.5 mmol). The mixturewas stirred for 2 hrs and concentrated in vacuo to remove dioxane. Waterwas added to the residue and solid crashed out. This solid was isolatedby filtration, washed with cold water thoroughly and dried in vacuumoven to give compound 249.4 as a tan solid. MS found for C₁₆H₁₄N₆OS as(M+H)⁺ 339.1.

Step 5: The above prepared compound 249.4 (150 mg, 0.44 mmol) wasdissolved in 6 mL NMP. To it was added MCPBA (65% pure, 140 mg, 0.53mmol). It was stirred at RT for 45 minutes to afford a mixture of thecorresponding sulfoxide and sulfone. To it then were added DIEA (230 μL,1.32 mmol) and then tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (81.2,142 mg, 0.66 mmol). The mixture was stirred for 2 hrs at 90° C. bath. Itwas then diluted with 150 mL EtOAc, washed with sat Na₂CO₃ and brine x2,dried over MgSO₄, filtered, and concentrated in vacuo. The residue wasthen exposed to 1:1 mixture of TFA and DCM for 1 hr. It was concentratedand subjected to reverse phase HPLC to isolate the title compound 249.MS found for C₂₁H₂₄N₈O as (M+H)⁺ 405.3. UV λ=249 nm. NMR (CD₃OD): δ 8.63(s, 1H), 8.59 (s, 1H), 8.57 (s, 1H), 8.24 (s, 1H), 8.01 (m, 1H), 7.27(m, 2H), 7.12 (dd, J=4.8, 4.8 Hz, 1H), 4.24 (m, 1H), 3.37 (m, 1H),1.64-1.21 (m, 8H) ppm.

Example 1212-((1R,2S)-2-aminocyclohexylamino)-4-(4-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline using a proceduresame as that described in Example 120. MS found for C₂₁H₂₄N₈O as (M+H)⁺405.3. UV λ=231, 314 nm. NMR (CD₃OD): δ 8.83 (d, J=4.8 Hz, 2H), 8.56 (s,1H), 8.45 (m, 2H), 7.81 (m, 2H), 7.35 (dd, J=5.2, 4.8 Hz, 1H), 4.44 (m,1H), 3.78 (m, 1H), 1.82-1.62 (m, 8H) ppm.

Example 1222-((1R,2S)-2-aminocyclohexylamino)-4-(3-(1-methyl-1H-pyrazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial 3-iodo-1-methylpyrazoleusing a procedure same as that described in Example 120. MS found forC₂₁H₂₆N₈O as (M+H)⁺ 407.3. UV λ=249 nm.

Example 1232-((1R,2S)-2-aminocyclohexylamino)-4-(3-(1-methyl-1H-pyrazol-4-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial 4-iodo-1-methylpyrazoleusing a procedure same as that described in Example 120. MS found forC₂₁H₂₆N₈O as (M+H)⁺ 407.3. UV λ=247 nm.

Example 1242-((1R,2S)-2-aminocyclohexylamino)-4-(3-(imidazo[1,2-a]pyridin-6-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial6-bromoimidazo[1,2-a]pyridine using a procedure same as that describedin Example 120. MS found for C₂₄H₂₆N₈O as (M+H)⁺ 443.3. UV λ=247 nm. NMR(CD₃OD): δ 9.17 (s, 1H), 8.57 (s, 1H), 8.31 (dd, J=9.6, 1.6 Hz, 1H),8.30 (d, J=1.6 Hz, 1H), 8.09 (d, J=2.4 Hz, 1H), 8.03 (d, J=9.2 Hz, 1H),7.95 (broad s, 1H), 7.87 (s, 1H), 7.63 (broad s, 2H), 4.35 (m, 1H), 3.66(m, 1H), 1.90-1.48 (m, 8H) ppm.

Example 1252-((1R,2S)-2-aminocyclohexylamino)-4-(4-(imidazo[1,2-a]pyridin-6-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using commercial6-bromoimidazo[1,2-a]pyridine and4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline using a proceduresame as that described in Example 120. MS found for C₂₄H₂₆N₈O as (M+H)⁺443.3. UV λ=245, 303 nm.

Example 1264-(1H-indazol-6-ylamino)-2-(1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

Step 1: To a stirring solution of carboxylic acid 202.1 (85 g, 540 mmol)in thionyl chloride (425 mL) was added pyridine (8.5 mL, 0.11 mmol),slowly. The reaction was stirred at 75° C. overnight at which time itwas concentrated and dried under vacuum to a light yellow powder. Thisyellow solid was slowly diluted with 750 mL of ethanol and refluxedovernight. The next day the reaction was determined to be complete byHPLC and then cooled in an ice bath and the solid filtered and washedwith diethyl ether affording ethyl ester 202.2 as an off-white powder(91 g, 87% for two steps). MS found for C₇H₈N₂O₄ as (M+H)⁺ 185.0.

Step 2: Ester 202.2 (22 g, 120 mmol) was dissolved in phosphorousoxychloride (60 mL, 600 mmol) and the mixture treated withN,N-diethylaniline (27 mL, 167 mmol) and the mixture heated to 105° C.until the reaction was determined to be complete by HPLC. It was thencooled to RT and slowly added to 1 L of crushed ice resulting in theformation of a beige precipitate which was collected by filtration anddried under vacuum affording dichloride 202.3 as a light yellow powder(22.5 g, 85%). ¹H NMR (DMSO-d₆, 400 MHz): δ 9.13 (s, 1H), 4.37 (q, 2H),1.32 (t, 3H).

Step 3: Dichloropyrimidine 202.3 (1.04 g, 4.7 mmol) was dissolved in NMP(30 mL) and stirred in ice bath. To it were added 6-aminoindazole 202.4(690 mg, 5.2 mmol) and then dropwise ethyldiisopropylamine (DIEA, 1.64mL, 9.4 mmol). The mixture was stirred for 40 minutes, and to it wasadded sodium thiomethoxide (660 mg, 9.4 mmol). The mixture was stirredfor overnight, diluted with ethyl acetate, washed with brine threetimes, and concentrated in vacuo to give crude compound 202.5 as a lightbrown solid in quantitative yield. MS found for C₁₅H₁₅N₅O₂S as (M+H)⁺330.1.

Step 4: Ethyl ester 202.5 (4.7 mmol) was dissolved in 60 mL THF. To itwere added lithium hydroxide hydrate (236 mg, 5.6 mmol) and 20 mL water.The mixture was stirred for overnight and to it was carefully added 1NHCl solution till pH reaching 2. The mixture was concentrated in vacuoto remove THF. White solid crashed out and was isolated using a Büchnerfunnel. It was washed with water and dried in vacuum oven to givecompound 202.6 (1.14 g, 81%) as a white solid. MS found for C₁₃H₁₁N₅O₂Sas (M+H)⁺302.1.

Step 5: Carboxylic acid 202.6 (1.14 g, 3.8 mmol) was dissolved in 30 mLDMF. To it were added EDC hydrochloride (1.09 g, 5.7 mmol) and HOBthydrate (770 mg, 5.7 mmol). The mixture was stirred at RT for 1 hour. Toit was then added ammonia (commercial 0.5N solution in dioxane, 22 mL,11.4 mmol). The mixture was stirred for 2 hours. It was thenconcentrated in vacuo and taken into water and ethyl acetate. Theorganic phase was separated and washed with brine four times. Theorganic phase was then dried over MgSO₄ and concentrated in vacuo toafford compound 202.7 as a light yellow solid (820 mg, 72%). MS foundfor C₁₃H₁₂N₆OS as (M+H)⁺ 301.1.

Step 6: Compound 202.7 (36 mg, 0.12 mmol) was dissolved in 3 mL NMP. Toit was added MCPBA (65% pure, 48 mg, 0.18 mmol). It was stirred at RTfor 30 minutes. To it then was added cis-1,2-diaminocyclohexane (71 μL,0.60 mmol). The mixture was stirred for 90 minutes at 90° C. bath. Thismixture was then subjected to preparative HPLC to isolate the racemictitle compound 202. MS found for C₁₈H₂₂N₈O as (M+H)⁺ 367.2. UV λ=245,300 nm.

Example 1272-((1R,2S)-2-aminocyclohexylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 6-aminobenzothiazole to replace6-aminoindazole 202.4. MS found for C₁₈H₂₁N₇OS as (M+H)⁺ 384.2. UVλ=241, 298 nm.

Example 1284-(1H-benzo[d][1,2,3]triazol-6-ylamino)-2-(1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide3

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 5/6-aminobenzotriazole to replace6-aminoindazole 202.4. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.2. UV λ=246,295 nm.

Example 1292-((1R,2S)-2-aminocyclohexylamino)-4-(2-methylbenzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 5-amino-2-methylbenzothiazole toreplace 6-aminoindazole 202.4. MS found for C₁₉H₂₃N₇OS as (M+H)⁺ 398.2.UV λ=246, 295 nm.

Example 1304-(1H-indol-6-ylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide6

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 6-aminoindole to replace6-aminoindazole 202.4. MS found for C₁₉H₂₃N₇O as (M+H)⁺ 366.2. UV λ=239,309 nm.

Example 1314-(1H-indazol-5-ylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 5-aminoindazole to replace6-aminoindazole 202.4. MS found for C₁₈H₂₂N₈O as (M+H)⁺ 367.2. UV λ=245,294 nm.

Example 1322-((1R,2S)-2-aminocyclohexylamino)-4-(quinolin-6-ylamino)pyrimidine-5-carboxamide

Step 1: Dichloropyrimidine 202.3 (500 mg, 2.3 mmol) was dissolved in NMP(20 mL) and stirred in ice bath. To it were added 6-aminoquinoline 208.1(390 mg, 2.7 mmol) and then dropwise ethyldiisopropylamine (DIEA, 0.72mL, 4.1 mmol). The mixture was stirred for 2 hours, diluted with ethylacetate, washed with brine three times, and concentrated in vacuo togive crude compound 208.2 as a light brown solid in quantitative yield.MS found for C₁₆H₁₃ClN₄O₂ as (M+H)⁺ 329.1.

Step 2: Ethyl ester 208.2 (2.3 mmol) was dissolved in 30 mL THF. To itwere added lithium hydroxide hydrate (193 mg, 4.6 mmol) and 6 mL water.The mixture was stirred for 7 hours and to it was carefully added 1N HClsolution till pH reaching 5. The mixture was concentrated in vacuo toremove THF and was extracted with ethyl acetate 5 times. The organicphases were combined, dried and concentrated in vacuo to give crude acid208.3. MS found for C₁₄H₉ClN₄O₂ as (M+H)⁺ 301.1.

Step 3: Carboxylic acid 208.3 (220 mg, 0.73 mmol) was dissolved in 18 mLNMP. To it were added EDC hydrochloride (210 mg, 1.1 mmol) and HOBthydrate (150 mg, 1.1 mmol). The mixture was stirred at RT for 1 hour. Toit was then added ammonia (commercial 0.5N solution in dioxane, 7.3 mL,3.65 mmol). The mixture was stirred for 2.5 hours. It was thenconcentrated in vacuo and taken into water and ethyl acetate. Theorganic phase was separated and washed with brine three times. Theorganic phase was then dried over MgSO₄ and concentrated in vacuo toafford compound 208.4 as a solid (180 mg, 62%). MS found for C₂₀H₁₄N₈O₂as (M+H)⁺ 399.1.

Step 6: Compound 208.4 (72 mg, 0.18 mmol) was dissolved in 3 mL NMP. Toit was added cis-1,2-diaminocyclohexane (100 μL, 0.90 mmol). The mixturewas stirred for 90 minutes at 90° C. bath. This mixture was thensubjected to preparative HPLC to isolate the racemic title compound 208.MS found for C₂₀H₂₃N₇O as (M+H)⁺ 378.2. UV λ=241, 283 nm.

Example 1334-(1H-benzo[d]imidazol-6-ylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 132 with tert-butyl6-amino-1H-benzo[d]imidazole-1-carboxylate to replace 6-aminoquinoline208.1. MS found for C₁₈H₂₂N₈O as (M+H)⁺ 367.2. UV λ=243, 294 nm.

Example 1342-((1R,2S)-2-aminocyclohexylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 5-aminobenzothiazole to replace6-aminoindazole 202.4. MS found for C₁₈H₂₁N₇OS as (M+H)⁺ 384.2. UVλ=246, 292 nm.

Example 1352-((1R,2S)-2-aminocyclohexylamino)-4-(imidazo[1,2-a]pyridin-6-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with imidazo[1,2-a]pyridin-6-amine toreplace 6-aminoindazole 202.4. MS found for C₁₈H₂₂N₈O as (M+H)⁺ 367.2.UV λ=250 nm.

Example 1362-((1R,2S)-2-aminocyclohexylamino)-4-(2,3-dihydrobenzo[b][1,4]dioxin-6-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 132 with 2,3-dihydrobenzo[b][1,4]dioxin-6-amineto replace 6-aminoquinoline 208.1. MS found for C₂₀H₂₃N₇O as (M+H)⁺385.2. UV λ=240, 294 nm. NMR (CD₃OD): δ 8.45 (s, 1H), 7.31 (d, J=2.4 Hz,1H), 6.91 (dd, J=8.4, 2.0 Hz, 1H), 6.85 (d, J=8.8 Hz, 1H), 4.27 (m, 5H),3.79 (m, 1H), 1.94-1.58 (m, 8H) ppm.

Example 1372-((1R,2S)-2-aminocyclohexylamino)-4-(quinoxalin-6-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 126 with 6-aminoquinoxaline to replace6-aminoindazole 202.4. MS found for C₁₉H₂₂N₈O as (M+H)⁺ 379.2. UV λ=242nm. NMR (CD₃OD): δ 8.87 (s, 1H), 8.82 (s, 1H), 8.74 (m, 1H), 8.61 (s,1H), 8.10 (d, J=8.8 Hz, 1H), 7.87 (d, J=8.4 Hz, 1H), 4.54 (m, 1H), 3.82(m, 1H), 1.99-1.62 (m, 8H) ppm.

Example 1382-((1R,2S)-2-aminocyclohexylamino)-4-(benzo[c][1,2,5]thiadiazol-5-ylamino)pyrimidine-5-carboxamide

The above racemic compound was prepared using the same synthetic schemedemonstrated in Example 132 with 2,1,3-benzothiadiazole-5-amine toreplace 6-aminoquinoline 208.1. MS found for C₁₇H₂₀N₈OS as (M+H)⁺ 385.2.UV λ=243 nm. NMR (CD₃OD): δ 8.73 (m, 1H), 8.60 (s, 1H), 7.96 (m, 1H),7.63 (dd, J=9.6, 2.0 Hz, 1H), 4.48 (m, 1H), 3.87 (m, 1H), 1.98-1.63 (m,8H) ppm.

Example 1392-((1R,2S)-2-aminocyclohexylamino)-4-(2-methylquinolin-6-ylamino)pyrimidine-5-carboxamide

Step 1: To a solution of Dichloropyrimidine 215.1 (700 mgs, 3.16 mmol)in acetonitrile (8 mL) was added a suspension of6-amino-2-methylquinoline (500 mgs, 3.16 mmol), diisopropylamine (0.61mL, 3.5 mmol) in acetonitrile (10 mL) at 0° C. Reaction mixture was thenslowly warmed to rt and stirred overnight. The reaction mixture was thendiluted with water and the precipitate collected by filtration affordingthe desired product 215.2 (964 mgs, 89%). MS found for C₁₇H₁₅ClN₄O₂ as(M+H)⁺ 343.1.

Step 2: Ethyl ester 215.2 (960 mgs, 2.81 mmol) was diluted with1,4-dioxane (7.5 mL) and ethanol (2 mL), followed by aqueous lithiumhydroxide (1.0 M, 2.8 mL, 2.8 mmol) and stirred at rt until all startingmaterial had been converted to the carboxylic acid. The reaction wasthen diluted with and acidified with 1N HCl (3.0 mL). The resultingsuspension was then filtered, washed with water and dried giving 870 mgsof the carboxylic acid 215.3 (98%). MS found for C₁₅H₁₁ClN₄O₂ as (M+H)⁺316.1.

Step 3: To carboxylic acid 215.3 (870 mgs, 2.76 mmol), EDC (792 mgs,4.14 mmol), HOBt (560 mgs, 4.14 mmol) in N,N-dimethylformamide (14 mL)was added ammonia (0.5 M in 1,4-dioxane, 14 mL, 6.9 mmol) and stirredovernight. The reaction mixture was then diluted with water (100 mL) andthe precipitate collected by filtration affording the desired product215.4 (1.10 g, 97%). MS found for C₂₁H₁₆N₈O₂ as (M+H)⁺ 413.1.

Step 4: A mixture of Benzotriazolyl ether 215.4 (75 mgs, 0.182 mmol),cis-1,2-diaminocyclohexane (25 mgs, 0.218 mmol), DIPEA (0.1 mL, 0.546mmol) in iso-propanol (3 mL) was heated in microwave (Emry's Optimizer)at 130° C. for 20 min.

The reaction mixture was then diluted with water and acetonitrile anddirectly purified by preparative HPLC affording the desired product,215, after lyophilization. MS found for C₂₁H₂₅N₇O as (M+H)⁺ 392.2.

Example 1402-((1R,2S)-2-aminocyclohexylamino)-4-(quinolin-5-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using 5-Aminoquinoline using a proceduresimilar to that described in Example 129. MS found for C₂₀H₂₃N₇O as(M+H)⁺ 378.3.

Example 1412-((1R,2S)-2-aminocyclohexylamino)-4-(2-methylquinolin-8-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using 8-Amino-2-methylquinoline using aprocedure similar to that described in Example 140. MS found forC₂₁H₂₅N₇O as (M+H)⁺392.3.

Example 1422-((1S,2R)-2-aminocyclohexylamino)-4-(quinolin-8-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using 8-Aminoquinoline using a proceduresimilar to that described in Example 139. MS found for C₂₀H₂₃N₇O as(M+H)⁺ 378.3.

Example 1432-((1R,2S)-2-aminocyclohexylamino)-4-(2-(morpholinomethyl)quinolin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using2-(morpholinomethyl)quinolin-6-amine (J. Med. Chem. 2006, 49, 7095)using a procedure similar to that described in Example 139. MS found forC₂₅H₃₂N₈O₂ as (M+H)⁺ 477.4.

Example 1442-((1R,2S)-2-aminocyclohexylamino)-4-(2,2-difluorobenzo[d][1,3]dioxol-5-ylamino)pyrimidine-5-carboxamide

Step 1: To a solution of Dichloropyrimidine 220.1 (700 mgs, 3.16 mmol)in acetonitrile (10 mL) was added a suspension of2,2-difluoro-5-aminobenzodioxole (549 mgs, 3.16 mmol), diisopropylamine(0.61 mL, 3.5 mmol) in acetonitrile (5 mL) at 0° C. Reaction mixture wasthen slowly warmed to rt and stirred overnight. The reaction mixture wasthen diluted with water (50 mL) and the precipitate collected byfiltration affording the desired product 220.2 (1.03 g, 91%). MS foundfor C₁₄H₁₀ClF₂N₃O₄ as (M+H)⁺ 358.1.

Step 2: Ethyl ester 220.2 (1.03 g, 2.9 mmol) was diluted with1,4-dioxane (7.5 mL) followed by aqueous lithium hydroxide (1.0 M, 2.9mL, 2.9 mmol) and stirred at rt until all starting material had beenconverted to the carboxylic acid. The reaction was then diluted withwater (20 mL) and acidified with 1N HCl (3.6 mL). The resultingsuspension was then filtered, washed with water and dried giving 950 mgsof the carboxylic acid 220.3 (99%). MS found for C₁₂H₆ClF₂N₃O₄ as (M+H)⁺330.0.

Step 3: To carboxylic acid 220.3 (950 mgs, 2.89 mmol), EDC (828 mgs,4.33 mmol), HOBt (663 mgs, 4.33 mmol) in N,N-dimethylformamide (14 mL)was added ammonia (0.5 M in 1,4-dioxane, 14 mL, 6.9 mmol) and stirredovernight. The reaction mixture was then diluted with water (60 mL) andthe precipitate collected by filtration affording the desired product220.4 (1.26 g, 99%). MS found for C₁₈H₁₁F₂N₇O₄ as (M+H)⁺ 428.2.

Step 4 and Step 5: A mixture of Benzotriazolyl ether 220.4 (75 mgs,0.176 mmol), tert-butyl (1S,2R)-2-aminocyclohexylcarbamate (45 mgs,0.211 mmol), DIPEA (0.1 mL, 0.530 mmol) in iso-propanol (3 mL) washeated in microwave (Emry's Optimizer) at 130° C. for 20 min. Thereaction mixture was concentrated and then was treated with 4.0M HCl indioxane (5.0 mL). After 1 h at rt, concentrate the reaction mixture anddiluted with water and acetonitrile and directly purified by preparativeHPLC affording the desired product 220, after lyophilization. MS foundfor C₁₈H₂₀F₂N₆O₃ as (M+H)⁺ 407.28.

Example 145

The following compounds were prepared using a procedure similar to thatdescribed in Example 143.

TABLE 1 Ex No Structure MW MS Name 146

381.44  382.35 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2- oxoindolin-5-ylamino)pyrimidine- 5-carboxamide 147

439.52  440.38 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2,2,4-trimethyl-3-oxo- 3,4-dihydro-2H- benzo[b][1,4]oxa- zin-6-ylamino)pyrimidine- 5-carboxamide 148

397.439 398.31 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(3-oxo-3,4-dihydro-2H- benzo[b][1,4]oxa- zin-7- ylamino)pyrimidine-5-carboxamide 149

409.494 410.36 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1- methyl-2-oxo-1,2,3,4- tetrahydroquinolin- 6- ylamino)pyrimidine- 5-carboxamide 150

402.462 403.32 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-cyanoquinolin-6- ylamino)pyrimidine- 5-carboxamide 151

395.467 396.35 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-oxo- 1,2,3,4-tetrahydroquinolin- 6- ylamino)pyrimidine- 5-carboxamide 152

393.451 394.14 2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-oxo- 1,2-dihydroquinolin-6- ylamino)pyrimidine- 5-carboxamide 153

411.466 412.5  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(4-methyl-3-oxo-3,4- dihydro-2H- benzo[b][1,4]oxa- zin-7-ylamino)pyrimidine- 5-carboxamide 154

397.439 398.5  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(3-oxo-3,4-dihydro-2H- benzo[b][1,4]oxa- zin-6- ylamino)pyrimidine-5-carboxamide 155

407.478 408.6  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1-methyl-2-oxo-1,2- dihydroquinolin-6- ylamino)pyrimidine- 5-carboxamide156

476.585 477.6  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-(morpholinomethyl) quinolin-6- ylamino)pyrimidine- 5-carboxamide 157

377.452 378.5  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(quinolin- 6-ylamino)pyrimidine- 5-carboxamide 158

380.46  381.1.  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1- methyl-1H-indazol-6- ylamino)pyrimidine- 5-carboxamide 159

380.46  381.1  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1- methyl-1H-indazol-5- ylamino)pyrimidine- 5-carboxamide 160

380.46  381.1  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1- methyl-1H-benzo[d]imidazol- 5- ylamino)pyrimidine- 5-carboxamide 161

462.56  463.3  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-morpholinoquinolin- 6- ylamino)pyrimidine- 5-carboxamide 162

397.49  398.2  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(4- methyl-3,4-dihydro-2H- benzo[b][1,4]oxa- zin-6- ylamino)pyrimidine- 5-carboxamide163

379.47  380.4  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(1-methyl-1H-indol- 5- ylamino)pyrimidine- 5-carboxamide 164

391.48  392.2  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(8-methylquinolin-4- ylamino)pyrimidine- 5-carboxamide 165

432.53  433.2  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(2-(cyclopropylamino) quinolin-6- ylamino)pyrimidine- 5-carboxamide 166

411.47  412.2  2-((1R,2S)-2- aminocyclohexyl- amino)-4-(4-methyl-3-oxo-3,4- dihydro-2H- benzo[b][1,4]oxa- zin-6-ylamino)pyrimidine- 5-carboxamide

Example 1672-((1R,2S)-2-aminocyclohexylamino)-4-(4′-(2-oxopyridin-1(2H)-yl)biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 MS found for C₂₈H₂₉N₇O₂ as (M+H)⁺ 496.5. UVλ=245 nm. NMR (CD₃OD): δ 8.51 (s, 1H), 8.12 (s, 1H), 7.79 (d, J=8.8 Hz,2H), 7.63 (m, 2H), 7.55-7.49 (m, 5H), 6.65 (m, 1H), 6.50 (m, 1H), 4.25(m, 1H), 3.58 (m, 1H), 2.54 (t, 2H), 1.84-1.40 (m, 8H) ppm.

Example 1682-((1R,2S)-2-aminocyclohexylamino)-4-(4′-(2-oxopiperidin-1-yl)biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 MS found for C₂₈H₃₃N₇O₂ as (M+H)⁺ 500.5. UVλ=249 nm. NMR (CD₃OD) δ 8.50 (s, 1H), 8.09 (s, 1H), 7.68 (d, J=8.8 Hz,2H), 7.49-7.43 (m, 3H), 7.36 (d, J=8.4 Hz, 2H), 4.21 (m, 1H), 3.71 (t,2H), 3.57 (m, 1H), 2.54 (t, 2H), 1.98 (m, 4H), 1.83-1.36 (m, 8H) ppm.

Example 1692-((1R,2S)-2-aminocyclohexylamino)-4-(3′-(2-oxopyridin-1(2H)-yl)biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120. MS found for C₂₈H₂₉N₇O₂ as (M+H)⁺ 496.5. UVλ=244 nm.

Example 170 2.2-((1R,2S)-2-aminocyclohexylamino)-4-(3′-(2-oxopiperidin-1-yl)biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120. MS found for C₂₈H₃₃N₇O₂ as (M+H)⁺ 500.5. UVλ=246 nm.

Example 1712-((1R,2S)-2-aminocyclohexylamino)-4-(4′-morpholinobiphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120. MS found for C₂₇H₃₃N₇O₂ as (M+H)⁺ 488.4. UVλ=247 nm.

Example 1722-((1R,2S)-2-aminocyclohexylamino)-4-(3′-morpholinobiphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 MS found for C₂₇H₃₃N₇O₂ as (M+H)⁺ 488.4. UVλ=246 nm.

Example 1732-((1R,2S)-2-aminocyclohexylamino)-4-(3′-morpholinobiphenyl-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120. MS found for C₂₇H₃₃N₇O₂ as (M+H)⁺ 488.4. UVλ=238, 309 nm.

Example 1742-((1R,2S)-2-aminocyclohexylamino)-4-(4′-morpholinobiphenyl-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120. MS found for C₂₇H₃₃N₇O₂ as (M+H)⁺ 488.4. UVλ=241, 314 nm.

Example 1752-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3,6-dihydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

Diisopropylamine (1.63 mL, 11.68 mmol) was dissolved in 10 mL dry THFand stirred in ice bath. To it was added n-buthyl lithium (2.5 M inhexane, 4.67 mL, 11.68 mmol) dropwise. The mixture was stirred for 20min, and sent to −78° C. bath. To it was added the solution of ketone257.1 (0.92 mL, 10 mmol) in 10 mL THF dropwise. The mixture was stirredfor 30 min. To it was added the solution ofN-phenylbis(trifluoromethanesulfonimide) (PhNTf₂, 4.17 g, 11.68 mmol) in10 mL THF. The mixture was moved to ice bath and stirred for overnight.It was concentrated in vacuo and subjected to silica flash column toisolate compound 257.2 using 20% ethyl acetate in hexane.

Compound 257.2 (920 mg, 4.0 mmol) was mixed with boronic acid 257.3 (550mg, 4.0 mmol), Pd(Ph₃P)₂Cl₂ (562 mg, 0.8 mmol), K₂CO₃ (1.1 g, 8.0 mmol)in 30 mL dioxane and 15 mL water. The mixture was degassed using argonstream for 3 min and stirred under argon at 85° C. for 3.5 h. Themixture was diluted with ethyl acetate, washed with brine x2, dried,concentrated and subjected to flash column to isolate aniline 257.4 (300mg) as white solid.

The title compound was prepared with aniline 257.4 using the samesynthetic scheme demonstrated in Example 120. MS found for C₂₂H₂₈N₆O₂ as(M+H)⁺ 409.4. UV λ=246 nm.

Example 1762-((1R,2S)-2-aminocyclohexylamino)-4-(3-(tetrahydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared from Example 175 using standardcatalysis hydrogenation by treating the solution of Example 175 inmethanol with 10% Pd/C under H₂ balloon for overnight. MS found forC₂₂H₃₀N₆O₂ as (M+H)⁺ 411.4. UV λ=241, 290 nm.

Example 1772-((1R,2S)-2-aminocyclohexylamino)-4-(4-(3,6-dihydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry shown forExample 175. MS found for C₂₂H₂₈N₆O₂ as (M+H)⁺ 409.4. UV λ=239, 309 nm.NMR (CD₃OD): δ 8.51 (s, 1H), 7.60 (d, J=7.2 Hz, 2H), 7.50 (d, J=8.0 Hz,2H), 6.23 (s, 1H), 4.36-4.30 (m, 3H), 3.94 (t, 2H), 3.72 (m, 1H), 2.53(m, 2H), 1.90-1.58 (m, 8H) ppm.

Example 1782-((1R,2S)-2-aminocyclohexylamino)-4-(4-(tetrahydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared from Example 175 using standardcatalysis hydrogenation by treating the solution of Example 175 inmethanol with 10% Pd/C under H₂ balloon for overnight. MS found forC₂₂H₃₀N₆O₂ as (M+H)⁺ 411.4. UV λ=243, 296 nm.

Example 1792-((1R,2S)-2-aminocyclohexylamino)-4-(4-(phenylsulfonyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with the corresponding aniline which wascommercially available. MS found for C₂₃H₂₆N₆O₃S as (M+H)⁺ 467.3. UVλ=232, 306 nm.

Example 1802-((1R,2S)-2-aminocyclohexylamino)-4-(4-phenoxyphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with the corresponding aniline which wascommercially available. MS found for C₂₃H₂₆N₆O₂ as (M+H)⁺ 419.3. UVλ=238, 290 nm.

Example 181a2-((1R,2S)-2-aminocyclohexylamino)-4-(4-(tetrahydro-2H-pyran-4-yloxy)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with the corresponding aniline which wascommercially available. MS found for C₂₂H₃₀N₆O₃ as (M+H)⁺ 427.3. UVλ=241, 291 nm.

Example 181b2-((1R,2S)-2-aminocyclohexylamino)-4-(4-(tetrahydro-2H-pyran-4-ylsulfonyl)phenylamino)pyrimidine-5-carboxamide

4-Nitrothiophenol (400 mg, 2.55 mmol) was dissolved in 10 mL DMF. To itwere added cesium carbonate (1.67 g, 5.1 mmol) and4-bromotetrahydropyran (0.84 g, 5.1 mmol). The mixture was stirred at50° C. for 90 min. It was diluted in ethyl acetate and washed with brinex3. The organic phase was dried, concentrated in vacuo to afford crudecompound 266.1. It was dissolved in 100 mL DCM. To it was adde MCPBA(1.98 g, 7.5 mmol) in small portions. The mixture was stirred for 30min, diluted with ethyl acetate, washed with sat sodium carbonatesolution and brine. The organic phase was dried, concentrated andsubjected to silica flash column to isolate compound 266.2 (320 mg, 47%for 2 steps) using 1:1 ethyl acetate and hexane.

Compound 266.2 (320 mg, 1.18 mmol) was dissolved in 150 mL ethylacetate. To it was added 200 mg 10% Pd/C. The mixture was stirred underH₂ balloon for overnight. The mixture was filtered through celite. Thecelite was thoroughly washed. The filtrate was concentrated in vacuo toafford aniline 266.3 (260 mg, 91%) as white solid.

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with aniline 266.3. MS found for C₂₂H₃₀N₆O₄Sas (M+H)⁺ 475.3. UV λ=250, 301 nm.

Example 1822-((1R,2S)-2-aminocyclohexylamino)-4-(4-(morpholinosulfonyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with the corresponding aniline which wascommercially available. MS found for C₂₁H₂₉N₇O₄S as (M+H)⁺ 476.4. UVλ=249, 300 nm.

Example 1832-((1R,2S)-2-aminocyclohexylamino)-4-(4-(morpholine-4-carbonyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with the corresponding aniline which wascommercially available. MS found for C₂₂H₂₉N₇O₃ as (M+H)⁺ 400.4. UVλ=244, 297 nm.

Example 1842-((1R,2S)-2-aminocyclohexylamino)-4-(3-(tetrahydro-2H-pyran-4-yloxy)phenylamino)pyrimidine-5-carboxamide

3-Nitrophenol (500 mg, 3.6 mmol) was dissolved in 15 mL DMF. To it wereadded cesium carbonate (2.35 g, 7.2 mmol) and then4-bromotetrahydropyran (2.4 g, 14.4 mmol). The mixture was stirred at70° C. for overnight. The reaction was only 30% completion. It wasdiluted with ethyl acetate, washed with water, sat sodium carbonate andbrine, dried over MgSO₄, filtered. To the filtrate containing crudecompound 269.1 was added 10% Pd/C 1.0 g. The mixture was stirred underH₂ balloon for overnight. The mixture was filtered, concentrated andpurified using silica flash column to afford aniline 269.2 (102 mg, 15%overall yield) as solid.

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with aniline 269.2. MS found for C₂₂H₃₀N₆O₃as (M+H)⁺ 427.4. UV λ=244, 287 nm.

Example 185 tert-butyl2-(4-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)acetate(racemate)

To a solution of 4-nitrophenol (3.00 g, 21.6 mmol) and K₂CO₃ (6.00 g,43.4 mmol) in DMF (50 mL), t-butyl bromoacetate (2.90 mL, 19.6 mmol) wasadded. The mixture was stirred at room temperature for 5 h. Water andEtOAc were added. The organic phase was separated, washed with water,then with aq. 1N NaOH and brine. It was dried over Na₂SO₄, concentratedin vacuo to give tert-butyl 2-(4-nitrophenoxy)acetate as a solid.

To the solution of the solid in MeOH (40 mL), Pd—C (10%, 410 mg) wasadded. The mixture was hydrogenated under balloon hydrogen for 20 h. Itwas then filtered through celite. The filtrate was concentrated in vacuoto give tert-butyl 2-(4-aminophenoxy)acetate.

The titled compound was synthesized analogously by using tert-butyl2-(4-aminophenoxy)acetate and cis-1,2-diaminocyclohexane. MS 457.2(M+H); UV 243.8, 290.0.

Example 1862-(4-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)aceticacid (racemate)

A solution of tert-butyl2-(4-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)acetateracemate (180 mg, 0.395 mmol) in TFA (4 mL) was stirred at roomtemperature for 2 h. It was then concentrated in vacuo. The residue waspurified by HPLC to give the titled compound (60 mg). MS 401.2 (M+H).

Example 187 tert-butyl2-(3-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)acetate(racemate)

The titled compound was synthesized analogously as compound tert-butyl2-(4-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)acetate(racemate), by using 3-nitrophenol in place of 4-nitrophenol. MS 457.2(M+H); UV 242.6, 290.0.

Example 1882-(3-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)aceticacid (racemate)

The titled compound was synthesized analogously as compound2-(4-(2-((1R,2S)-2-aminocyclohexylamino)-5-carbamoylpyrimidin-4-ylamino)phenoxy)aceticacid (racemate). MS 401.2 (M+H); UV 242.6, 290 nM.

Example 1892-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-pyrazol-4-ylamino)pyrimidine-5-carboxamide

The titled compound was synthesized analogously by using1-methyl-1H-pyrazol-4-amine. MS 331.4 (M+H). UV 241.2, 292.1 nm

Example 1902-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-pyrrol-3-ylamino)pyrimidine-5-carboxamide

To sodium hydride (300 mg, 60% in mineral oil, 7.50 mmol) in a flask,which was washed with hexane twice, a solution of 3-nitropyrrole (500mg, 4.46 mmol) and iodomethane (0.560 mL, 8.97 mmol) in DMF (4 mL) wasadded. Hydrogen gas evolved. The mixture was then stirred at roomtemperature for 20 h. Water and EtOAc were added. Organic phase wasseparated, washed with water, dried over Na₂SO₄, concentrated in vacuoto give 1-methyl-3-nitro-1H-pyrrole (494 mg).

A mixture of 1-methyl-3-nitro-1H-pyrrole (488 mg, 3.87 mmol) and Pd—C(10%, 110 mg) in MeOH (15 mL) (containing 8 drops 0f 6N HCl) washydrogenated under ballon hydrogen for 20 h. The mixture was filteredthrough celite. To the filtrate, 4N HCl in dioxane (2 mL) was added. Thesolution was concentrated in vacuo to give 1-methyl-1H-pyrrol-3-aminehydrochloride as a solid (514 mg).

The titled compound was then synthesized analogously by using1-methyl-1H-pyrrol-3-amine hydrochloride. MS 330.3 (M+H); UV 237.6,313.1 nm.

Example 1912-((1R,2S)-2-aminocyclohexylamino)-4-(1-phenyl-1H-pyrazol-4-ylamino)pyrimidine-5-carboxamide

A mixture of 4-nitropyrazole (270 mg, 2.38 mmol), iodobenzene (485 mg,2.38 mmol), 8-hydroxyquinoline (60 mg, 0.41 mmol) and K2CO3 (600 mg,4.34 mmol) in DMSO (3 mL) was degassed with argon before being chargedwith CuI (45 mg, 0.23 mmol). The mixture in a sealed tube was heated at130 C for 20 h. Water was added to induce precipitation. The precipitatewas collected to give 4-nitro-1-phenyl-1H-pyrazole (454 mg).

A mixture of 4-nitro-1-phenyl-1H-pyrazole (440 mg, 2.32 mmol) and SnCl2dihydrate (2.18 g, 9.66 mmol) in EtOAc (15 mL) was stirred at 80 C for 3h. Aqueous 1N NaOH was added to bring pH to 12. The mixture was filteredthrough celite. The organic phase was separated, dried over Na₂SO₄,concentrated in vacuo. The residue was purified by HPLC to give1-phenyl-1H-pyrazol-4-amine as a solid (173 mg).

The titled compound was then synthesized analogously by using1-phenyl-1H-pyrazol-4-amine. MS 393.4 (M+H); UV 240.0, 303.2.

Example 1922-((1R,2S)-2-aminocyclohexylamino)-4-(4-methylthiophen-2-ylamino)pyrimidine-5-carboxamide

A solution of 4-methyl thiophene-2-carboxylic acid (1.42 g, 10.0 mmol),triethylamine (1.50 mL, 10.8 mmol) and diphenyl phosphoryl azide (2.15mL, 10.0 mmol) in tBuOH (20 mL) was stirred at reflux for 5 h. tBuOH wasremoved in vacuo. Et2O and water were added. The organic phase waswashed with 5% NaHCO₃, dried over Na₂SO₄, concentrated in vacuo. Theresidue was purified by a silica gel column, which was eluted with 0-10%EtOAc in hexane to give tert-butyl 4-methylthiophen-2-ylcarbamate as asolid (0.880 g).

A solution of tert-butyl 4-methylthiophen-2-ylcarbamate (0.880 g, 4.13mmol) in CH₂Cl₂ (8 mL) and TFA (6 mL) wad stirred at room temperaturefor 3 h. Solvents was removed in vacuo to give 4-methylthiophen-2-amineas trifluoroacetic acid salt (0.920 g).

The titled compound was then synthesized analogously by using4-methylthiophen-2-amine. MS 347.3 (M+H); UV 244.9, 326.1 nm.

Example 1932-((1R,2S)-2-aminocyclohexylamino)-4-(5-methylthiophen-2-ylamino)pyrimidine-5-carboxamide

A solution of 5-methyl thiophene-2-carboxylic acid (1.42 g, 10.0 mmol),triethylamine (1.50 mL, 10.8 mmol) and diphenyl phosphoryl azide (2.15mL, 10.0 mmol) in tBuOH (20 mL) was stirred at reflux for 5 h. tBuOH wasremoved in vacuo. Et2O and water were added. The organic phase waswashed with 5% NaHCO₃, then with 1N NaOH, and filtered. The filtrate wasdried over Na₂SO₄, concentrated in vacuo to give tert-butyl5-methylthiophen-2-ylcarbamate as a solid (0.825 g).

A solution of tert-butyl 5-methylthiophen-2-ylcarbamate (0.825 g, 3.87mmol) in CH₂Cl₂ (10 mL) and TFA (6 mL) wad stirred at room temperaturefor 20 h. Solvents was removed in vacuo to give 5-methylthiophen-2-amineas trifluoroacetic acid salt (0.870 g).

The titled compound was then synthesized analogously by using5-methylthiophen-2-amine. MS 347.3 (M+H); UV 247.3, 325.6 nm.

Example 1942-((1R,2S)-2-aminocyclohexylamino)-4-(1-phenyl-1H-pyrrol-3-ylamino)pyrimidine-5-carboxamide

A mixture of 3-nitropyrrole (270 mg, 2.41 mmol), iodobenzene (0.267 mL,2.39 mmol), 8-hydroxyquinoline (60 mg, 0.41 mmol) and K2CO3 (600 mg,4.34 mmol) in DMSO (3 mL) was degassed with argon before being chargedwith CuI (45 mg, 0.23 mmol). The mixture in a sealed tube was heated at130 C for 20 h. Water and EtOAc were added. The organic phase wasseparated, washed with 1N HCl, then with 5% NaHCO₃, dried over Na₂SO₄,concentrated in vacuo to give 3-nitro-1-phenyl-1H-pyrrole (410 mg).

A mixture of 3-nitro-1-phenyl-1H-pyrrole (410 mg, 2.18 mmol) and SnCl2dihydrate (2.00 g, 8.86 mmol) in EtOAc (15 mL) was stirred at 80 C for 3h. Aqueous 1N NaOH was added to bring pH to 12. The mixture was filteredthrough celite. The organic phase was separated, washed with 5% NaHCO₃,dried over Na₂SO₄, concentrated in vacuo to give1-phenyl-1H-pyrrol-3-amine as a solid (323 mg).

The titled compound was then synthesized analogously by using1-phenyl-1H-pyrrol-3-amine. MS 392.4 (M+H); UV 240.0, 314.3 nM.

Example 1952-((1R,2S)-2-aminocyclohexylamino)-4-(4-(6-(dimethylamino)pyridine-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 45 with 6-(dimethylamino)pyridin-3-ylboronicacid to replace boronic acid 116.1. MS found for C₂₄H₃₀N₈O as (M+H)⁺447.3. UV: λ=244.0, 316.4

Example 1962-((1R,2S)-2-aminocyclohexylamino)-4-(3,5-difluoro-4-morpholinophenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 3,5-difluoro-4-morpholinoaniline toreplace aniline 72.4. MS found for C₂₁H₂₇F₂N₇O₂ as (M+H)⁺ 448.1. UV:λ=240.4, 304.5.

Example 1972-((1R,2S)-2-aminocyclohexylamino)-4-(biphenyl-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with biphenyl-4-amine to replace aniline 72.4.MS found for C₂₃H₂₆N₆O as (M+H)⁺ 403.4. UV: λ=239.3, 308.7.

Example 1982-((1R,2S)-2-aminocyclohexylamino)-4-(3-(isoxazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 3-(isoxazol-3-yl)aniline to replaceaniline 72.4. MS found for C₂₀H₂₃N₇O₂ as (M+H)⁺ 394.4. UV: λ=243.6,286.5.

Example 1994-(4-(4H-1,2,4-triazol-4-ylphenylamino)-2-(1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 4-(4H-1,2,4-triazol-4-yl)aniline toreplace aniline 74.1. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV:λ=245.2, 293.8.

Example 2004-(3-(4H-1,2,4-triazol-4-ylphenylamino)-2-(1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 3 with 3-(4H-1,2,4-triazol-4-yl)aniline toreplace aniline 74.1. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV:λ=239.9.

Example 2012-((1R,2S)-2-aminocyclohexylamino)-4-(4-(isoxazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 4-(isoxazol-3-yl)aniline to replaceaniline 74.1. MS found for C₂₀H₂₃N₇O₂ as (M+H)⁺ 394.4. UV: λ=244.0,303.3

Example 2022-((1R,2S)-2-aminocyclohexylamino)-4-(3-(5-methylisoxazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 3-(5-methylisoxazol-3-yl)aniline toreplace aniline 74.1. MS found for C₂₁H₂₅N₇O₂ as (M+H)⁺ 408.4. UV:λ=243.6, 287.1.

Example 203 Methyl2-((1S,2R)-2-(5-carbamoyl-4-(m-tolylamino)pyrimidin-2-ylamino)cyclohexylamino)acetate

MS found for C₂₁H₂₈N₆O₃ as (M+H)⁺ 413.1. UV: λ=241.4, 288.8.

Example 2042-((1S,2R)-2-(5-carbamoyl-4-(m-tolylamino)pyrimidin-2-ylamino)cyclohexylamino)aceticacid

MS found for C₂₀H₂₆N₆O₃ as (M+H)⁺ 399.2. UV: λ=240.5, 287.8.

Example 2052-((1S,2R)-2-aminocyclohexylamino)-4-(4-methoxy-3-methylphenylamino)pyrimidin-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 4-methoxy-3-methylaniline to replaceaniline 72.4. MS found for C₁₉H₂₆N₆O₂ as (M+H)⁺ 371.2. UV: λ=238.1,292.6.

Example 2062-((1S,2R)-2-aminocyclohexylamino)-4-(3,4-dimethoxyphenylamino)pyrimidin-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 3,4-dimethoxyaniline to replace aniline72.4. MS found for C₁₉H₂₆N₆O₃ as (M+H)⁺ 387.1. UV: λ=236.9, 286.6.

Example 2072-((1S,2R)-2-aminocyclohexylamino)-4-(3-phenoxyphenylamino)pyrimidin-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with 3-phenoxyaniline to replace aniline 72.4.MS found for C₂₃H₂₆N₆O₂ as (M+H)⁺ 419.3. UV: λ=240.4, 292.6.

Example 2082-((1R,2S)-2-aminocyclohexylamino)-4-(biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 1 with biphenyl-3-amine to replace aniline 72.4.MS found for C₂₃H₂₆N₆O as (M+H)⁺ 403.4. UV: λ=246.3

Example 2092-((1R,2S)-2-aminocyclohexylamino)-4-(naphthalen-1-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with naphthalen-1-amine to replace aniline208.1. MS found for C₂₁H₂₄N₆O as (M+H)⁺ 377.1.

Example 2102-((1R,2S)-2-aminocyclohexylamino)-4-(6-methoxynaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 6-methoxynaphthalen-2-amine to replaceaniline 208.1. MS found for C₂₂H₂₆N₆O₂ as (M+H)⁺ 407.2. UV: λ=227.5,319.9.

Example 211a2-((1R,2S)-2-aminocyclohexylamino)-4-(6-fluoronaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 6-fluoronaphthalen-2-amine to replaceaniline 208.1. MS found for C₂₁H₂₃FN₆O as (M+H)⁺ 395.1. UV: λ=212.2,244.0, 306.8.

Example 211b2-((1R,2S)-2-aminocyclohexylamino)-4-(6-carbamoylnaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 6-carbamoylnaphthalen-2-amine toreplace aniline 208.1. MS found for C₂₂H₂₅N₇O₂ as (M+H)⁺ 420.2. UV:λ=223.9, 318.8.

Example 2122-((1R,2S)-2-aminocyclohexylamino)-4-(6-(methylcarbamoyl)naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 6-(methylcarbamoyl)naphthalen-2-amineto replace aniline 208.1. MS found for C₂₃H₂₇N₇O₂ as (M+H)⁺ 434.3. UV:λ=219.2, 235.7, 318.8.

Example 2132-((1R,2S)-2-aminocyclohexylamino)-4-(6-(dimethylcarbamoyl)naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 6-(dimethylcarbamoyl)naphthalen-2-amineto replace aniline 208.1. MS found for C₂₄H₂₉N₇O₂ as (M+H)⁺ 448.2. UV:λ=218.0, 314.0.

Example 2142-((1R,2S)-2-aminocyclohexylamino)-4-(4-chloronaphthalen-1-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 4-chloronaphthalenyl-1-amine to replaceaniline 208.1. MS found for C₂₁H₂₃ClN₆O as (M+H)⁺ 411.2, 413.1 (Clpattern). UV: λ=223.9, 293.8.

Example 2152-((1R,2S)-2-aminocyclohexylamino)-4-(6-bromonaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with 4-bromonaphthalenyl-2-amine to replaceaniline 208.1. MS found for C₂₁H₂₃BrN₆O as (M+H)⁺ 455.1, 457.1 (Brpattern).

Example 2172-((1R,2S)-2-aminocyclohexylamino)-4-(6-(morpholine-4-carbonyl)naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 152 with6-(morpholine-4-carbonyl)naphthalenyl-2-amine to replace aniline 208.1.MS found for C₂₆H₃₁N₇O₃ as (M+H)⁺ 490.4. UV: λ=220.4, 315.2.

Example 2182-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

Step 1: To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (328mg, 1.48 mmol) and 1-methyl-1H-indol-4-amine (260 mg, 1.78 mmol) inCH₃CN (6 mL) at room temperature, DIEA (0.4 mL, 2.22 mmol) was added.The mixture was stirred at room temperature for 24 h. Water (15 mL) wasadded to induce precipitation. The precipitate was collected, dried onvacuum to give ethyl 2-chloro-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxylate as a solid.

Step 2: To a solution of ethyl 2-chloro-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxylate (crude from step 1) in THF (4 mL), aq. 1N LiOH(2.25 mL, 2.25 mmol) was added. The mixture was stirred at roomtemperature overnight. Upon acidification of the mixture with 1N HCl,white solids precipitated out, which were collected, and dried on vacuumto give 2-chloro-4-(1-methyl-1H-indol-4-ylamino) pyrimidine-5-carboxylicacid (325 mg). MS 303.3, 305.3 (M+H, Cl pattern)

Step 3: To a solution of2-chloro-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxylic acid(325 mg, 1.08 mmol) and HOBt (198 mg, 1.29 mmol) in DMF (4 mL), EDC (248mg, 1.29 mmol) was added. The mixture was stirred at room temperaturefor 1.5 h. Ammonia (0.5 M in dioxane, 8.00 mL, and 4.00 mmol) was added.It was stirred at room temperature overnight. Water and EtOAc wereadded. The organic phase was separated, washed with 1 N HCl, then with5% NaHCO₃, dried over Na₂SO₄, concentrated in vacuo to give2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide(378 mg). MS 401.4 (M+H)

Step 4: To a solid of2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide(100 mg, 0.25 mmol) in vial was added tert butyl (1S,2R)-2-aminocyclohexylcarbamate (0.3 M solution in NMP, 1.25 mL, 0.375mmol) and DIPEA (0.09 mL, 0.5 mmol). It was heated at 80° C. for 2 h,cooled and purified by preparative HPLC to give2-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide(21 mg). MS found for C₂₀H₂₅N₇O as (M+H)⁺ 380.4. UV: λ=219.2, 241.6,336.7.

Example 219a 4-(1H-indol-4-ylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1H-indol-4-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₃N₇O as (M+H)⁺ 366.3. UV:λ=216.7, 239.9, 330.3.

Example 219b 4-(1H-indazol-4-ylamino)-2-((1R,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1H-indazol-4-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₁₈H₂₂N₈O as (M+H)⁺ 367.4. UV:λ=205.8, 240.5, 314.3

Example 219c2-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1-methyl-1H-indazol-4-ylamine toreplace 1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₄N₈O as (M+H)⁺381.4. UV: λ=.

Example 2202-((1R,2S)-2-aminocyclohexylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 2-methyl-2H-indazol-4-ylamine toreplace 1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₄N₈O as (M+H)⁺381.5. UV: λ=210.6, 243.0, 329.1.

Example 2212-((1R,2S)-2-aminocyclohexylamino)-4-(2-methylbenzo[d]oxazol-7-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 2-methylbenzo[d]oxazol-7-ylamine toreplace 1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₃N₇O₂ as (M+H)⁺382.4. UV: λ=238.1.

Example 2222-((1R,2S)-2-aminocyclohexylamino)-4-(benzo[c][1,2,5]thiadiazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with benzo[c][1,2,5]thiadiazol-4-ylamine toreplace 1-methyl-1H-indol-4-ylamine. MS found for C₁₇H₂₀N₈OS as (M+H)⁺385.3. UV: λ=234.5, 298.5, 315.2.

Example 2232-((1R,2S)-2-aminocyclohexylamino)-4-(quinoxalin-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with quinoxalinyl-5-amine to replace1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₂N₈OS as (M+H)⁺ 379.3.UV: λ=203.4, 245.4.

Example 2242-((1R,2S)-2-aminocyclohexylamino)-4-(benzo[d]thiazol-7-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with benzo[d]thiazol-7-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₁₈H₂₁N₇OS as (M+H)⁺ 384.3.UV: λ=205.1, 242.8, 290.2.

Example 2252-((1R,2S)-2-aminocyclohexylamino)-4-(1-methyl-1H-benzo[d]imidazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1-methyl-1H-benzo[d]imidazol-4-ylamineto replace 1-methyl-1H-indol-4-ylamine. MS found for C₁₉H₂₄N₈O as (M+H)⁺381.4. UV: λ=202.8, 239.3.

Example 2262-((1R,2S)-2-aminocyclohexylamino)-4-(2-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 2-methyl-1H-indol-4-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₂₀H₂₅N₇O as (M+H)⁺ 380.4. UV:λ=220.4, 239.3, 336.7.

Example 2272-((1R,2S)-2-aminocyclohexylamino)-4-(2-phenyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 2-phenyl-1H-indol-4-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₂₅H₂₇N₇O as (M+H)⁺ 442.5. UV:λ=241.6, 293.8.

Example 2282-((1R,2S)-2-aminocyclohexylamino)-4-(1,2-dimethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1,2-dimethyl-1H-indol-4-ylamine toreplace 1-methyl-1H-indol-4-ylamine. MS found for C₂₁H₂₇N₇O as (M+H)⁺394.4. UV: λ=222.8, 242.8.

Example 2292-((1R,2S)-2-aminocyclohexylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-c]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 120 with2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-amine to replace1-methyl-1H-indol-4-ylamine. MS found for C₂₂H₂₇N₇O as (M+H)⁺ 406.5. UV:λ=222.8, 241.6.

Example 2302-((1R,2S)-2-aminocyclohexylamino)-4-(benzo-Misoxazol-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with benzo-Misoxazol-5-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₁₈H₂₁N₇O₂ as (M+H)⁺ 368.4.UV: λ=203.9, 236.9, 294.9.

Example 2312-((1R,2S)-2-aminocyclohexylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218 with 1-ethyl-1H-indol-4-ylamine to replace1-methyl-1H-indol-4-ylamine. MS found for C₂₁H₂₇N₇O as (M+H)⁺ 394.4. UV:λ=220.2, 242.6.

Example 2322-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3,4-dimethoxyphenyl)phenylamino)pyrimidine-5-carboxamide

Potassium carbonate (1.81 g, 13.1 mmol) was dissolved in a 2:1 mixtureof dioxane/water (45 mL). This solution was added to3,4-dimethoxyphenylboronic acid (300.1; 799 mg, 4.39 mmol) [CAS122775-35-3] and 3-bromoaniline (300.2; 756 mg, 4.39 mmol) [CAS591-19-5]. The resulting solution was degassed with argon for 5 minutes.Bis(triphenylphosphine) palladium(II) dichloride (598 mg, 0.85 mmol)[CAS 13965-03-2] was added and the reaction was stirred for 3 h at 85°C. in a sealed tube. The reaction was cooled and diluted with EtOAc (300mL), washed with brine (3×100 mL), dried over MgSO₄, filtered, andconcentrated. The resulting residue was subjected to flashchromatography (gradient of 20% to 70% EtOAc in hexanes) which resultedin 360 mg of 300.3 (36%).

To 300.3 (360 mg, 1.57 mmol) in DMF (15 mL) was added4-chloro-2-(methylthio)-5-pyrimidinecarboxylic acid ethyl ester (365 mg,1.57 mmol) [CAS 5909-24-0] and DIEA (562 μL, 3.14 mmol). The reactionmixture was stirred for 3 h at 80° C. in a sealed tube. The reaction wascooled, water was added, and a precipitate formed. The precipitate wasfiltered, washed with cold water, and dried to give 300.4 inquantitative yield.

The resulting solid (667 mg, 1.57 mmol) was dissolved in THF (10 mL). Tothis was added LiOH (188 mg, 7.85 mmol) in H₂O (5 mL). The reaction wasstirred for 30 min and was acidified to pH-3 with 1 M HCl. The THF wasremoved in vacuo and ice-cold water was added to the reaction mixture.The resulting solid was filtered, washed with water, and dried to give497 mg (88%) of 300.5.

Carboxylic acid 300.5 (460 mg, 1.15 mmol) was dissolved in 10 mL DMF. Toit were added EDC hydrochloride (328 mg, 1.72 mmol) and HOBt hydrate(232 mg, 1.72 mmol). The mixture was stirred at RT for 2 h. Ammonia(commercial 0.5N solution in dioxane, 7 mL, 3.5 mmol) was added and themixture and was stirred for 1 h. The dioxane was removed by in vacuo andice-cold water was added to the reaction mixture. The resulting solidwas filtered, washed with water, and dried to give 445 mg (97%) of300.6.

Compound 300.6 (70 mg, 0.18 mmol) was dissolved in 4 mL NMP. To it wasadded mCPBA (70% minimum purity, 50 mg, 0.21 mmol) which was stirred atRT for 30 minutes.

To it were then added a solution of tert-butyl(1S,2R)-2-aminocyclohexylcarbamate (0.3 M, 0.9 mL, 0.27 mmol) and DIEA(94 μL, 0.54 mmol). The mixture was stirred for 3 h at 90° C. in asealed tube. The mixture was cooled, diluted with EtOAc, washed with asaturated Na₂CO₃ aqueous solution, water, and brine. The organic phasewas dried over MgSO₄ and concentrated afford a crude intermediate whichwas stirred in a 1:1 mixture of TFA and DCM at RT for 30 min. Thereaction mixture was concentrated in vacuo and subjected to reversephase preparative HPLC to isolate title compound. MS found forC₂₅H₃₀N₆O₃ as (M+H)⁺ 463.3. UV λ=210, 243 nm. δ 1.40-1.83 (m, 8H),3.60-3.68 (m, 1H), 3.88 (s, 3H), 3.92 (s, 3H), 4.43-4.51 (m, 1H),7.60-7.66 (m, 1H), 7.90 (d, 1H), 8.04 (s, 2H), 8.52-8.58 (m, 1H), 8.62(s, 1H).

Example 2332-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as shown forexample 232. However, 4-methoxyphenylboronic acid [CAS 5720-07-0] wasused instead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MSfound for C₂₄H₂₈N₆O₂ as (M+H)⁺ 433.4. UV λ=248 nm. δ 1.25-1.80 (m, 8H),3.58-3.64 (m, 1H), 3.83 (s, 3H), 4.20-4.28 (m, 1H), 7.02 (d, 2H),7.36-7.48 (m, 3H), 7.59 (d, 2H), 8.03 (br s, 1H), 8.55 (s, 1H)

Example 2342-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,3,4-trimethoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as example 232.However, 3,4,5-trimethoxyphenylboronic acid [CAS 182163-96-8] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₆H₃₂N₆O₄ as (M+H)⁺ 493.4. UV λ=245 nm. δ 1.20-1.80 (m, 8H),3.61-3.68 (m, 1H), 3.80 (s, 3H), 3.92 (s, 6H), 4.15-4.22 (m, 1H), 6.92(s, 2H), 7.45 (br s, 3H), 8.05 (br s, 1H), 8.55 (s, 1H).

Example 2352-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3-methoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as example 232.However, 3-methoxyphenylboronic acid [CAS 10365-98-7] was used insteadof 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS found forC₂₄H₂₈N₆O₂ as (M+H)⁺ 433.3. UV λ=241 nm.

Example 2362-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,3-dihydrobenzofuran-5-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as example 232.However, (2,3-dihydrobenzo[b]furan-5-yl)boronic acid [CAS 227305-69-3]was used instead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3].MS found for C₂₅H₂₈N₆O₂ as (M+H)⁺ 445.3. UV λ=245, 276 nm.

Example 2372-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2-methoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as example 232.However, 2-methoxyphenylboronic acid [CAS 5720-06-9] was used instead of3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS found forC₂₄H₂₈N₆O₂ as (M+H)⁺ 433.3. UV λ=244, 288 nm.

Example 2382-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-fluorophenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 4-fluorophenylboronic acid [CAS 1765-93-1] was used instead of3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS found forC₂₃H₂₅FN₆O as (M+H)⁺421.3. UV λ=245 nm. δ 1.35-1.85 (m, 8H), 3.58-3.65(m, 1H), 4.25-4.31 (m, 1H), 7.09-7.18 (m, 1H), 7.40-7.54 (m, 6H), 8.11(br s, 1H), 8.53 (s, 1H).

Example 2392-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3-fluorophenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-fluorophenylboronic acid [CAS 768-35-4] was used instead of3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS found forC₂₃H₂₅FN₆O as (M+H)⁺ 421.3. UV λ=242 nm.

Example 2402-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,5-dimethoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 2,5-dimethoxyphenylboronic acid [CAS 107099-99-0] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₅H₃₀N₆O₃ as (M+H)⁺ 463.3. UV λ=246 nm.

Example 2412-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,4-dimethoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 2,4-dimethoxyphenylboronic acid [CAS 133730-34-4] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₅H₃₀N₆O₃ as (M+H)⁺ 463.3. UV λ=245, 288 nm.

Example 2422-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3,4-difluorophenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3,4-difluorophenylboronic acid [CAS 168267-41-2] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₃H₂₄F₂N₆O as (M+H)⁺ 439.3. UV 2=245 nm.

Example 2432-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,3-dimethoxyphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 2,3-dimethoxyphenylboronic acid [CAS 40972-86-9] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₅H₃₀N₆O₃ as (M+H)⁺ 463.3. UV λ=244, 288 nm.

Example 2442-((1R,2S)-2-aminocyclohexylamino)-4-(3-(benzo[d][1,3]dioxol-4-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline [CAS210907-84-9] and 4-bromo-1,3-benzodioxole [CAS 6698-13-1] were used ascoupling partners. MS found for C₂₄H₂₆N₆O₃ as (M+H)⁺ 447.4. UV λ=242 nm.

Example 2452-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline [CAS210907-84-9] and 6-iodobenzodioxane [CAS 57744-67-9] were used ascoupling partners. MS found for C₂₅H₂₈N₆O₃ as (M+H)⁺ 461.4. UV 2=211,243 nm. δ 1.35-1.85 (m, 8H), 3.58-3.65 (m, 1H), 4.30-4.38 (m, 5H), 6.95(d, 1H), 7.10-7.18 (m, 2H), 7.30-7.36 (m, 1H), 7.38-7.45 (m, 2H), 8.14(br s, 1H), 8.50 (s, 1H)

Example 2462-((1R,2S)-2-aminocyclohexylamino)-4-(4-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline [CAS214360-73-3] and 6-iodobenzodioxane [CAS 57744-67-9] were used ascoupling partners. MS found for C₂₅H₂₈N₆O₃ as (M+H)⁺ 461.4. UV λ=242,316 nm. δ 1.50-1.95 (m, 8H), 3.70-3.78 (m, 1H), 4.25 (s, 4H), 4.34-4.41(m, 1H), 6.90 (d, 1H), 7.08-7.15 (m, 2H), 7.58-7.68 (m, 4H), 8.50 (s,1H).

Example 2472-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methyl-3,4-dihydro-2H-benzo[b][1,4]oxazin-7-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline [CAS210907-84-9] and 7-bromo-4-methyl-3,4-dihydro-2H-benzo[1,4]oxazine [CAS154264-95-6] were used as coupling partners. MS found for C₂₆H₃₁N₇O₂ as(M+H)⁺ 474.4. UV λ=216, 238, 303 nm.

Example 2482-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-piperidinylphenyl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and N-(4-bromophenyl)piperidine [CAS 22148-20-5] ascoupling partners. The resulting aniline was reacted with 72.3 andsubjected to the subsequent chemistry shown in example 13 to yield thetitle compound. MS found for C₂₈H₃₅N₇O as (M+H)⁺ 486.5. UV λ=252 nm.

Example 2492-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3-piperidinylphenyl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and N-(3-bromophenyl)piperidine [CAS 84964-24-9] ascoupling partners. The resulting aniline was reacted with 72.3 andsubjected to the subsequent chemistry shown in example 13 to yield thetitle compound. MS found for C₂₈H₃₅N₇O as (M+H)⁺ 486.5. UV λ=247 nm.

Example 2502-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-pyrrolidinylphenyl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and N-(4-bromophenyl)pyrrolidine [CAS 22090-26-2] ascoupling partners. The resulting aniline was reacted with 72.3 andsubjected to the subsequent chemistry shown in example 13 to yield thetitle compound. MS found for C₂₇H₃₃N₇O as (M+H)⁺ 472.5. UV λ=246 nm.

Example 2512-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-pyrrolidin-2-oxo-ylphenyl)phenylamino)pyrimidine-5-carboxamide

This example was a byproduct in the formation of example 250. MS foundfor C₂₇H₃₁N₇O₂ as (M+H)⁺ 486.5. UV λ=248, 282 nm.

Example 2522-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3-pyrrolidin-2-oxo-ylphenyl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and N-(3-bromophenyl)pyrrolidine [CAS 219928-13-9] ascoupling partners. The resulting aniline was reacted with 72.3 andsubjected to the subsequent chemistry shown in example 13 to yield thetitle compound. MS found for C₂₇H₃₁N₇O₂ as (M+H)⁺ 486.5. UV λ=245 nm.

Example 2532-((1R,2S)-2-aminocyclohexylamino)-4-(3-(benzo[d][1,3]dioxol-5-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 1,3-benzodioxole-5-boronic acid [CAS 94839-07-3] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₄H₂₆N₆O₃ as (M+H)⁺447.3. UV λ=239, 274, 296 nm. δ 1.35-1.85 (m,8H), 3.58-3.65 (m, 1H), 4.25-4.31 (m, 1H), 6.00 (s, 2H) 6.93 (d, 1H),7.10-7.18 (m, 2H), 7.37-7.50 (m, 3H), 8.08 (br s, 1H), 8.53 (s, 1H)

Example 2542-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2-methylsulfonylphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 2-methylsulfonylphenylboronic acid [CAS 330804-03-0] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₄H₂₈N₆O₃S as (M+H)⁺ 481.4. UV λ=240, 285 nm.

Example 2552-((1R,2S)-2-aminocyclohexylamino)-4-(3-(3-methylsulfonylphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-methylsulfonylphenylboronic acid [373384-18-0] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₄H₂₈N₆O₃S as (M+H)⁺ 481.4. UV λ=246 nm.

Example 2562-((1R,2S)-2-aminocyclohexylamino)-4-(3-(4-methylsulfonylphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 4-methylsulfonylphenylboronic acid [149104-88-1] was usedinstead of 3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. MS foundfor C₂₄H₂₈N₆O₃S as (M+H)⁺481.4. UV λ=250 nm. δ 1.25-1.85 (m, 8H), 3.18(s, 3H), 3.58-3.65 (m, 1H), 4.21-4.28 (m, 1H), 7.52-7.62 (m, 3H) 7.93(d, 2H), 8.07 (d, 2H), 8.17 (br s, 1H), 8.57 (s, 1H)

Example 2572-((1R,2S)-2-aminocyclohexylamino)-4-(4-(3-methylsulfonylphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 3-methylsulfonylphenylboronic acid [373384-18-0] and4-iodoaniline [CAS 540-37-4] were used as coupling partners. MS foundfor C₂₄H₂₈N₆O₃S as (M+H)⁺ 481.4. UV λ=235, 308 nm.

Example 259 2-((1R,2S)-2-aminocyclohexylamino)-4-(4-(4-methylsulfonylphenyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 4-methylsulfonylphenylboronic acid [149104-88-1] and4-iodoaniline [CAS 540-37-4] were used as coupling partners. MS foundfor C₂₄H₂₈N₆O₃S as (M+H)⁺ 481.3. UV λ=239, 313 nm.

Example 2602-((1R,2S)-2-aminocyclohexylamino)-4-(3-(quinolin-4-yl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and 4-bromoquinoline [CAS 3964-04-3] as couplingpartners. The resulting aniline was reacted with 72.3 and subjected tothe subsequent chemistry shown in example 13 to yield the titlecompound. MS found for C₂₆H₂₇N₇O as (M+H)⁺ 454.4. UV λ=239, 302 nm.

Example 2612-((1R,2S)-2-aminocyclohexylamino)-4-(3-(quinolin-8-yl)phenylamino)pyrimidine-5-carboxamide

The initial biarylaniline was prepared utilizing the same chemistry asExample 232 using 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)aniline[CAS 210907-84-9] and 8-bromoquinoline [CAS 16567-18-3] as couplingpartners. The resulting aniline was reacted with 72.3 and subjected tothe subsequent chemistry shown in example 13 to yield the titlecompound. MS found for C₂₆H₂₇N₇O as (M+H)⁺ 454.4. UV λ=240, 301 nm.

Example 2622-((1R,2S)-2-aminocyclohexylamino)-4-(3-(quinolin-5-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the same chemistry as Example 232.However, 5-quinolinylboronic acid [CAS 355386-94-6] was used instead of3,4-dimethoxyphenylboronic acid [CAS 122775-35-3]. The resulting anilinewas reacted with 72.3 and subjected to the subsequent chemistry shown inexample 13 to yield the title compound. MS found for C₂₆H₂₇N₇O as (M+H)⁺454.5. UV λ=241, 302 nm.

Example 2642-((1R,2S)-2-aminocyclohexylamino)-4-(3-dibenzofuranamino)pyrimidine-5-carboxamide

This compound was prepared reacting commercially available3-aminodibenzofuran [CAS 4106-66-5] with 300.4 and DIEA. Subsequentreactions utilizing chemistry shown in Example 232 provided the titlecompound. MS found for C₂₃H₂₄N₆O₂ as (M+H)⁺ 417.4. UV λ=212, 236, 322nm.

Example 2652-((1R,2S)-2-aminocyclohexylamino)-4-(2-dibenzofuranamino)pyrimidine-5-carboxamide

2-Bromodibenzofuran (331.1; 942 mg, 3.81 mmol) [CAS 86-76-0], tert-butylcarbamate (670 mg, 5.72 mmol) [CAS 4248-19-5], and Cs₂CO₃ were added todegassed dioxane. Xantphos (330 mg, 0.51 mmol) [CAS 161265-03-8] andPd₂(dba)₃ (175 mg, 0.19 mmol) [CAS 51364-51-3] were subsequently added.The reaction was heated under argon at 85° C. for 18 h. The reaction wascooled and subjected to flash chromatography to give 331.2 which wasdissolved in 4 N HCl/dioxane. Reaction stirred for 12 h and then wasconcentrated in vacuo to yield 331.3 (200 mg). This aniline was reactedwith 72.3 and subjected to the subsequent chemistry shown in example 120to yield the title compound to yield the title compound. MS found forC₂₃H₂₄N₆O₂ as (M+H)⁺ 417.4. UV λ=210, 244, 289 nm.

Example 2662-((1R,2S)-2-aminocyclohexylamino)-4-(9-methyl-9H-carbazol-3-ylamino)pyrimidine-5-carboxamide

To 3-bromocarbazole (332.1) (530 mg, 2.15 mmol) [CAS 1592-95-6] in DMF(˜20 mL) at 0° C. was added NaH in DMF (˜5 mL). The reaction was warmedto RT and then heated at 60° C. for 1 h. The solution was allowed tocool to RT and iodomethane was added dropwise. The reaction was heatedat 12 h 60° C. in a sealed tube. The reaction mixture was cooled dilutedwith EtOAc and washed with water (4 times), brine, dried over MgSO4 andconcentrated to yield 332.2 (550 mg).

Intermediate 332.2 was subjected to chemistry seen in example 265 toyield 332.3. This aniline was reacted with 72.3 and subjected to thesubsequent chemistry shown in example 13 to yield the title compound toyield the title compound. MS found for C₂₃H₂₄N₆O₂ as (M+H)⁺ 430.4. UVλ=238, 296 nm.

Example 2674-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1R,2R)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 87 with commercially available (1R,2R)-cyclohexane-1,2-diamineand DIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=250 nm.

Example 2684-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1S,2S)-2-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 87 with commercially available (1S,2S)-cyclohexane-1,2-diamineand DIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=250 nm.

Example 2694-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,3S)-3-aminocyclohexylamino)pyrimidine-5-carboxamide(racemic)

The title racemic compound was prepared using the same chemistry shownfor Example 87 with commercially available cis-cyclohexane-1,3-diamineand DIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=252 nm.

Example 2704-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1R,3R)-3-aminocyclohexylamino)pyrimidine-5-carboxamide(racemic)

The title racemic compound was prepared using the same chemistry shownfor Example 87 with commercially available trans-cyclohexane-1,3-diamineand DIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=252 nm.

Example 2714-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1S,4S)-4-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 87 with commercially available cis-cyclohexane-1,4-diamine andDIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=252 nm.

Example 2724-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,4R)-4-aminocyclohexylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417 with commercially available trans-cyclohexane-1,4-diamineand DIEA. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV λ=252 nm.

Example 2734-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1S,2S)-2-aminocyclopentylamino)pyrimidine-5-carboxamide(racemic)

The title racemic compound was prepared using the same chemistry shownfor Example 417 with commercially availabletrans-cyclopentane-1,2-diamine and DIEA. MS found for C₁₈H₂₁N₉O as(M+H)⁺ 380.4. UV λ=251 nm.

Example 2742-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2H-1,2,3-triazol-2-yl)-5-(trifluoromethyl)phenylamino)pyrimidine-5-carboxamide

A mixture of 3-bromo-5-(trifluoromethyl)aniline 335.1 (1.08 g, 4.50mmol) [CAS 54962-75-3, 1H-1,2,3-triazole 335.2 (1.04 mL, 18.0 mmol),K₃PO₄ (1.91 g, 9.00 mmol), CuI (428 mg, 2.25 mmol),N,N′-dimethylethylenediamine (0.29 mL, 2.70 mmol) in 12 mL dioxane and 3mL DMSO was stirred in a sealed tube at 120° C. for 5 days. A mixture of335.3 and 335.4 (in ˜1.4:1 ratio) was obtained. The mixture was dilutedwith EtOAc (250 mL), washed with water, brine, dried over MgSO₄,filtered, and was concentrated in vacuo. The crude mixture was subjectedto flash column chromatography to isolate 335.3.

Aniline 335.3 was reacted with 300.4 according to reaction conditionsoutlined in Example 232. The chemistry of the scheme was completed toyield the title compound. MS found for C₂₀H₂₂F₃N₉O as (M+H)⁺ 462.3. UVλ=259 nm. δ 1.50-2.00 (m, 8H), 3.61-3.68 (m, 1H), 4.80-4.88 (m, 1H),7.80 (s, 1H), 8.05 (s, 2H), 8.15 (s, 1H), 8.60 (s, 1H), 9.05 (s, 1H)

Example 2752-((1R,2S)-2-aminocyclohexylamino)-4-(3-methyl-4-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

A mixture of 4-fluoro-3-methylnitrobenzene 337.1 (819 mg, 5.28 mmoL)[CAS 455-88-9], 1H-1,2,3-triazole 335.2 (1.23 mL 21.1 mmol) [CAS288-36-8], and cesium carbonate (3.44 g, 10.6 mmol) in 25 mL dry NMP wasstirred in a sealed tube at 120° C. for 24 h. It was cooled, dilutedwith 400 mL ethyl acetate and washed with water. The aqueous phase wasfurther extracted with EtOAc (2×200 mL). The combined organic phase waswashed with water and brine, dried over MgSO₄ and filtered to yield asolution of crude products 337.2 and 337.3 in ˜1:1 ratio. This solutionwas concentrated to 100 mL and a catalytic amount of 10% Pd/C was added.To this suspension was mounted a hydrogen balloon for overnightstirring. The mixture was filtered through celite and concentrated invacuo to afford crude anilines 337.4 and 337.5. MS found for C₉H₁₀N₄ as(M+H)⁺ 175.1. The two anilines were purified using flash column.

Aniline 337.4 was reacted with 300.4 and was subjected to the chemistryshown in Example 232 to yield the title compound. MS found for C₂₀H₂₅N₉Oas (M+H)⁺ 408.4. UV λ=239, 296 nm. δ 1.5-2.0 (m, 8H), 2.38 (s, 3H),3.65-3.75 (m, 1H), 4.35-4.45 (m, 1H), 7.52-7.62 (m, 1H), 7.65-7.75 (m,2H), 7.95 (s, 2H), 8.55 (s, 1H).

Example 2762-((1R,2S)-2-aminocyclohexylamino)-4-(3-methyl-4-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing the chemistry in example 275.Specifically, pyrazole [CAS 288-13-1] and 337.1 were reacted to form3-methyl-4-(1H-pyrazol-1-yl)benzenamine. This aniline was then reactedwith 300.4 according to conditions outlined in Example 232. Thechemistry of the scheme was completed to yield the title compound. MSfound for C₂₁H₂₆N₈O as (M+H)⁺ 407.5. UV λ=238, 296 nm. δ 1.5-2.0 (m,8H), 2.25 (s, 3H), 3.60-3.70 (m, 1H), 4.35-4.45 (m, 1H), 6.55 (s, 1H),7.40 (d, 1H), 7.60-7.68 (m, 2H), 7.75 (br s, 1H), 7.82 (br s, 1H), 8.55(s, 1H).

Example 2772-((1R,2S)-2-aminocyclohexylamino)-4-(3-(2H-1,2,3-triazol-2-yl)-4-(trifluoromethyl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared utilizing a modified procedure from thechemistry in example 275. Notably,2-Fluoro-4-nitro-1-trifluoromethylbenzene [CAS 69411-67-2] was usedinstead of 2-Fluoro-1-methyl-4-nitrobenzene [CAS 1427-07-2], and theinitial displacement reaction was stirred at 45° C. for 3 h. Afterhydrogenation, the resulting aniline was reacted with 300.4 according toconditions outlined in Example 232. The chemistry of the scheme wascompleted to yield the title compound. MS found for C₁₇H₁₈F₃N₉O as(M+H)⁺ 462.3. UV λ=245, 295 nm. δ 1.50-1.95 (m, 8H), 3.58-3.64 (m, 1H),4.43-4.51 (m, 1H), 7.60-7.66 (m, 1H), 7.90 (d, 1H), 8.04 (s, 2H),8.52-8.58 (m, 1H), 8.62 (s, 1H).

Example 2782-((1R,2S)-2-aminocyclohexylamino)-4-(3-(1H-1,2,3-triazol-1-yl)-4-(trifluoromethyl)phenylamino)pyrimidine-5-carboxamide

The 1-substituted triazole precursor of the title compound was formed inthe reaction described in example 277. After hydrogenation, theresulting aniline was reacted with 300.4 according to conditionsoutlined in Example 232. The chemistry of the scheme was completed toyield the title compound. MS found for C₁₇H₁₈F₃N₉O as (M+H)⁺ 462.3. UVλ=245, 295 nm. δ 1.45-1.85 (m, 8H), 3.58-3.64 (m, 1H), 4.38-4.46 (m,1H), 7.63-7.71 (m, 1H), 7.89-7.97 (m, 2H), 8.34 (br s, 1H), 8.49-8.56(m, 1H), 8.62 (s, 1H).

Example 2792-((1R,2S)-2-aminocyclohexylamino)-4-(3-vinylphenylamino)pyrimidine-5-carboxamide

Commerically available 3-vinylaniline was reacted with 72.3 andsubjected to the subsequent chemistry shown in example 87 to yield thetitle compound. MS found for C₁₉H₂₄N₆O as (M+H)⁺ 353.4. UV λ=244 nm.

Example 2802-(cis-2-aminocyclohexylamino)-4-(4-vinylphenylamino)pyrimidine-5-carboxamide

The title compound was synthesized utilizing the chemistry in Example11. However, instead of aniline 72.4, 4-vinylaniline [CAS 1520-21-4] wasutilized. MS found for C₁₉H₂₄N₆O as (M+H)⁺ 353.4. UV λ=236, 310 nm.

Example 2812-(cis-2-amino-cis-3-methylcyclohexylamino)-4-(3-methylphenylamino)pyrimidine-5-carboxamide

This compound was synthesized utilizing the same chemistry shown inExample 302.

MS found for C₁₉H₂₆N₆O as (M+H)⁺ 355.4. UV λ=242, 289 nm. δ 1.00 (d,3H), 1.20-1.95 (m, 7H), 2.40 (s, 3H), 3.18 (s, 3H), 3.72-3.80 (m, 1H),4.02-4.10 (m, 1H), 7.07-7.14 (m, 1H) 7.25-7.35 (m, 2H), 7.41-7.48 (m,1H), 8.54 (s, 1H).

Example 2822-(cis-2-amino-cis-3-methylcyclohexylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

This compound was synthesized utilizing the same chemistry shown inExample 303. MS found for C₂₀H₂₈N₆O as (M+H)⁺ 369.4. UV λ=240, 290 nm. δ1.00 (d, 3H), 1.20-1.95 (m, 7H), 2.38 (s, 6H), 3.70-3.78 (m, 1H),4.04-4.12 (m, 1H), 6.95 (s, 1H), 7.19-7.25 (m, 2H), 8.50 (s, 1H).

Example 2832-(cis-2-amino-cis-3-methylcyclohexylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesized utilizing the same chemistry shown inExample 298. MS found for C₂₀H₂₅N₉O as (M+H)⁺ 408.4. UV λ=250 nm. δ0.75-0.85 (m, 3H), 1.20-1.95 (m, 7H), 3.62-3.70 (m, 1H), 4.20-4.26 (m,1H), 7.38-7.42 (m, 1H), 7.55 (t, 1H), 7.89-7.94 (m, 1H), 7.97 (s, 2H),8.56 (s, 1H), 8.66-8.70 (m, 1H).

Example 2842-(cis-6-aminocyclohex-3-enylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesized utilizing the same chemistry shown inExample 298.

MS found for C₁₉H₂₁N₉O as (M+H)⁺ 392.3. UV λ=250 nm.

Example 2852-((1R,2S)-2-aminocyclohexylamino)-4-(3-fluoro-4-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The 3-fluoro-4-(2H-1,2,3-triazole) aniline intermediate was prepared byreacting 1H-1,2,3-triazole and 4-bromo-3-fluoroaniline [CAS 656-65-5]according to example 92. This intermediate was then reacted with 300.4according to the reaction conditions outlined in Example 232. Thechemistry of the scheme was completed to yield the title compound. MSfound for C₁₉H₂₂FN₉O as (M+H)⁺ 412.4. UV λ=242, 305 nm. δ 1.5-2.0 (m,8H), 3.75-3.85 (m, 1H), 4.40-4.50 (m, 1H), 7.45 (d, 1H), 7.75-7.85 (m,1H), 7.95-8.1 (m, 3H), 8.60 (s, 1H).

Example 286 2-((1SR,2S)-2-aminocyclohexylamino)-4-(phenethylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 9. MS found for C₁₉H₂₆N₆O as (M+H)⁺ 355.4. UV:λ=232.2.

Example 2872-((1S,2R)-1-amino-1,2,3,4-tetrahydronaphthalen-2-ylamino)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The above compound was prepared using similar procedure as described. MSfound for C₁₈H₂₂N₆O as (M+H)⁺ 339.2.

Example 2882-((1R,2S)-2-amino-1,2,3,4-tetrahydronaphthalen-1-ylamino)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The above compound was prepared using similar procedure as described. MSfound for C₁₈H₂₂N₆O as (M+H)⁺ 339.2.

Example 2892-((1R,2S)-2-aminocyclohexylamino)-4-(naphthalen-2-ylamino)pyrimidine-5-carboxamide

Step 1: To a solution of Dichloropyrimidine 215.1 (700 mgs, 3.16 mmol)in acetonitrile (8 mL) was added a suspension of 6-amino-naphthylene(3.16 mmol), diisopropylamine (0.61 mL, 3.5 mmol) in acetonitrile (10mL) at 0° C. Reaction mixture was then slowly warmed to rt and stirredovernight. The reaction mixture was then diluted with water and theprecipitate collected by filtration affording the desired product 215.2.

Step 2: Ethyl ester 215.2 (960 mgs, 2.81 mmol) was diluted with1,4-dioxane (7.5 mL) and ethanol (2 mL), followed by aqueous lithiumhydroxide (1.0 M, 2.8 mL, 2.8 mmol) and stirred at rt until all startingmaterial had been converted to the carboxylic acid. The reaction wasthen diluted with and acidified with 1N HCl (3.0 mL). The resultingsuspension was then filtered, washed with water and dried giving 870 mgsof the carboxylic acid 215.3.

Step 3: To carboxylic acid 215.3 (870 mgs, 2.76 mmol), EDC (792 mgs,4.14 mmol), HOBt (560 mgs, 4.14 mmol) in N,N-dimethylformamide (14 mL)was added ammonia (0.5 M in 1,4-dioxane, 14 mL, 6.9 mmol) and stirredovernight. The reaction mixture was then diluted with water (100 mL) andthe precipitate collected by filtration affording the desired product215.4.

Step 4: A mixture of Benzotriazolyl ether 215.4 (75 mgs, 0.182 mmol),cis-1,2-diaminocyclohexane (25 mgs, 0.218 mmol), DIPEA (0.1 mL, 0.546mmol) in iso-propanol (3 mL) was heated in microwave (Emry's Optimizer)at 130° C. for 20 min.

The reaction mixture was then diluted with water and acetonitrile anddirectly purified by preparative HPLC affording the desired product,215, after lyophilization. MS found for C₂₁H₂₂N₆O as (M+H)⁺ 377.3.

Example 2902-((1S,2R)-1-amino-1,2,3,4-tetrahydronaphthalen-2-ylamino)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The above compound was prepared using a similar procedure as describedherein. MS found for C₁₈H₂₂N₆O as (M+H)⁺ 339.2.

Example 2912-((1R,2S)-2-amino-1,2,3,4-tetrahydronaphthalen-1-ylamino)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The above compound was prepared using similar procedure as described. MSfound for C₁₈H₂₂N₆O as (M+H)⁺ 339.2.

Example 2932-((1S,2R)-2-(5-carbamoyl-4-(m-tolylamino)pyrimidin-2-ylamino)cyclohexylamino)aceticacid

The above compound was prepared using a similar procedure as describedherein. MS found for C₂₀H₂₆N₆O₃ as (M+H)⁺ 399.2. UV: λ=240.5, 212.2,287.8.

Example 2942-(cis-4-aminocyclohexylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 271. MS found for C₂₀H₂₅N₇O as (M+H)⁺ 380.4. UV:λ=245.4.

Example 2952-(cis-2-aminocyclopentylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was synthesized utilizing the same chemistry shown inExample 87.

MS found for C₁₈H₂₁N₉O as (M+H)⁺ 380.4. UV λ=249 nm.

Example 2962-(cis-2-aminocyclopentylamino)-4-(3-methylphenylamino)pyrimidine-5-carboxamide

Compound 344 was synthesized utilizing similar chemistry as that foundin example 1. The benzotrizole containing precursor 344.1 was reactedwith cis-diamine 343.4 to give the title compound. MS found forC₁₇H₂₂N₆O as (M+H)⁺ 327.4. UV λ=240, 287 nm.

Example 2972-(cis-2-aminocyclopentylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

Compound 345 was synthesized utilizing similar chemistry as that foundin example 1. The benzotrizole containing precursor 345.1 was reactedwith cis-diamine 343.4 to give the title compound. MS found forC₁₈H₂₄N₆O as (M+H)⁺ 341.4. UV 2=217, 239, 290 nm.

Example 2984-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)pyrimidine-5-carboxamide

Step 1: 3,4-Di-O-acetyl-D-arabinal [CAS 3945-17-3] (179.1) was made bystandard literature method or bought as commercially available chemicalreagent. It (5.0 g, 25 mmol) was dissolved in 100 mL methanol. To it wasadded 1.0 gram of 10% Pd/C and the mixture was stirred under a hydrogenballoon for overnight to give compound 179.2. The mixture was filteredto remove Pd/C. The filtrate was treated with sodium methoxide (5.4 g,100 mmol) at RT for overnight. It was quenched with 6N HCl (20 mL) inice bath. The mixture was concentrated in vacuo to dryness. The residuewas treated with 500 mL ethyl acetate and vigorously stirred at 45° C.for 30 min. The slurry was filtrered using a fine-grade Büchner funnel.The filtrate was concentrated in vacuo and pumped overnight to afforddiol 179.3 (2.82 g, 95%).

Step 2: Diol 179.3 (205 mg, 1.74 mmol) was dissolved in 10 mL dry DCM.To it were added pyridine (0.35 mL, 4.34 mmol) and in the ice bath Tf₂O(0.63 mL, 3.74 mmol). The reaction was monitored by TLC (stain: ammoniummolybdate (12.5 g), Ce(IV) sulfate (5 g), 10% H₂SO₄ 500 mL; heated usingstrong heat from a hot heat gun after dipping). The reaction was over in15 min. To the mixture were added 24 mL DMF and 3 mL HMPA. Then sodiumazide (2.28 g, 35 mmol) was added. The mixture was then stirred at 50°C. for 3 h (reaction was complete in 1.5 h by TLC). The mixture wasdiluted with ethyl acetate, filtered, washed with brine. The organicphase was dried, concentrated and subjected to silic flash column toisolate diazide 179.4 (220 mg, 75%).

Step 3: Diazide 179.4 (220 mg, 1.3 mmol) was dissolved in 40 mL ethylacetate. It was treated with 10% Pd/C under H₂ balloon for overnight.The mixture was filtered through celite and concentrated in vacuo tooffer crude diamine 179.5.

Step 4: Intermediate 179.6 was made using aniline 156.6 (Example 66)using the same chemistry shown in Example 1. MS found for C₁₄H₁₃N₇OS as(M+H)⁺ 328.2.

Step 5: Methylthio pyrimidine 179.6 (105 mg, 0.32 mmol) was dissolved in2 mL NMP and treated with MCPBA (102 mg, 0.38 mmol) at RT for 45 min. Toit were added DIEA (226 μL, 1.3 mmol) and diamine 179.5 made in Step 3.The mixture was stirred 2 h at 90° C. for 2 h. Two major products foundwere 179 and 179.7 at 1.58 min and 1.55 min by reverse phase analyticalHPLC (5 min run) in ratio of 1.7:1 in favor of 179. Compound 179 wasisolated from the mixture using reverse phase prep HPLC. MS found forC₁₈H₂₁N₉O₂ as (M+H)⁺ 396.3. UV λ=249 nm. NMR (CD₃OD): δ 8.66 (s, 1H),8.52 (s, 1H), 7.93 (s, 2H), 7.82 (m, 1H), 7.47 (m, 1H), 7.27 (m, 1H),4.40 (m, 1H), 4.00 (m, 1H), 3.78 (m, 1H), 3.72 (m, 1H), 3.55 (m, 1H),3.42 (m, 1H), 2.00 (m, 1H), 1.86 (m, 1H) ppm.

Example 2994-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(3S,4S)-4-aminotetrahydro-2H-pyran-3-ylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298 using 3,4-Di-O-acetyl-L-arabinal [CAS 3945-18-4], commerciallyavailable, to replace 3,4-Di-O-acetyl-D-arabinal [CAS 3945-17-3]. Thetitle compound was the minor product isolated from the reaction mixturein the final step. MS found for C₁₈H₂₁N₉O₂ as (M+H)⁺ 396.3. UV λ=250 nm.NMR (CD₃OD): δ 8.73 (s, 1H), 8.58 (s, 1H), 8.09 (s, 2H), 7.90 (m, 1H),7.54 (m, 1H), 7.34 (m, 1H), 4.50 (m, 1H), 4.06 (m, 1H), 3.83 (m, 1H),3.76 (m, 1H), 3.63 (m, 1H), 3.60 (m, 1H), 3.43 (m, 1H), 2.08 (m, 1H),1.92 (m, 1H) ppm.

Example 3004-(3-(1H-pyrazol-1-yl)phenylamino)-2-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298. MS found for C₁₉H₂₂N₈O₂ as (M+H)⁺ 395.4. UV λ=246 nm.

Example 3012-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298. MS found for C₂₀H₂₂N₈O₂ as (M+H)⁺ 407.4. UV λ=249 nm.

Example 3022-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298. MS found for C₁₇H₂₂N₆O₂ as (M+H)⁺ 343.3. UV λ=240, 288 nm.

Example 3032-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298. MS found for C₁₈H₂₄N₆O₂ as (M+H)⁺ 357.4. UV λ=240, 290 nm.

Example 3042-((3R,4R)-3-aminotetrahydro-2H-pyran-4-ylamino)-4-(3-methoxyphenylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example298. MS found for C₁₇H₂₂N₆O₃ as (M+H)⁺ 359.4. UV 2=240, 283 nm. NMR(CD₃OD): δ 8.46 (s, 1H), 7.29 (m, 1H), 7.26 (m, 1H), 6.99 (m, 1H), 6.74(m, 1H), 4.25 (m, 1H), 4.03 (m, 1H), 3.89 (m, 1H), 3.79 (m, 1H), 3.77(s, 3H), 3.62 (m, 1H), 3.55 (m, 1H), 2.02 (m, 1H), 1.82 (m, 1H) ppm.

Example 3052-((3S,4S)-4-aminotetrahydro-2H-pyran-3-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

This compound was prepared using the same chemistry shown for Example299. MS found for C₁₇H₂₂N₆O₂ as (M+H)⁺ 343.3. UV λ=241, 287 nm. NMR(CD₃OD): δ 8.53 (s, 1H), 7.47 (m, 1H), 7.37 (m, 1H), 7.28 (m, 1H), 7.02(m, 1H), 4.50 (m, 1H), 4.05 (m, 1H), 4.01 (m, 1H), 3.73-3.68 (m, 2H),3.57 (m, 1H), 2.38 (s, 3H), 2.05 (m, 1H), 1.87 (m, 1H) ppm.

Example 3062-((1R,2R)-2-amino-3,3-difluorocyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic)

Step 1: 2-Cyclohexen-1-ol (5.0 mL, 50 mmol) was dissolved in 100 mL DCM.To it were added pyridine (12.2 mL, 150 mmol) and TBDMSC1 (12.1 g, 80mmol). The mixture was stirred at RT for overnight. The mixture wasconcentrated in vacuo to remove pyridine. The residue was taken into 300mL ethyl acetate and 200 mL water. The organic phase was separated,washed with brine, dried, concentrated and purified using silica flashcolumn using 15% ethyl acetate in hexane. Compound 187.1 was thus got asoil (10.1 g, 94%).

Step 2: Compound 187.1 (10.1 g, 47.7 mmol) was dissolved in 70 mLacetonitrile. To it was added NMO (50% water solution, 22.5 mL, 95.4mmol) and OsO₄ (4% water solution, 9.1 mL, 1.43 mmol). The mixture wasstirred overnight at RT. It was diluted with 600 mL ethyl acetate,washed with brine, sat sodium bicarbonate, dried, concentrated andpurified by silica flash column using 60% ethyl acetate in hexane togive compound 187.2 (10.8 g, 92%) as oil.

Step 3: Diol 187.2 (10.8 g, 43.9 mmol) was dissolved in 200 mL DCM. Inica bath, to it were added pyridine (17.7 mL, 219 mmol) and dropwiseMSCl (8.2 mL, 105 mmol). The mixture was stirred for overnight at RT. Itwas dilute with 500 mL DCM, washed with brine x3, dried, concentrated,purified using silica flash column with 45% ethyl acetate in hexane togive compound 187.3 (16.8 g, 95%) as oil.

Step 4: Compound 187.3 (15.7 g, 39 mmol) was dissolved in 80 mL DMF and20 mL HMPA. To it was added sodium azide (16.5 g, 253 mmol). The mixturewas stirred at 100° C. for overnight (24 h). The mixture was dilutedwith 800 mL ethyl acetate, washed with water and brine x2, dried,concentrated and purified by silica flash column using 10% ethyl acetatein hexane to isolate diazide 187.4 (3.16 g, 27%) as oil.

Step 5: Compound 187.4 (1.21 g, 4.1 mmol) was dissolved in 60 mL dryTHF. To it was added TBAF (1.0 M in THF, 8.2 mL, 8.2 mmol). The mixturewas stirred for 2 h. It was quenched with methanol, concentrated invacuo and subjected to silica flash column using 40-50% ethyl acetate inhexane to isolated compound 187.5 (854 mg, 99%) as oil.

Step 6: Compound 187.5 (450 mg, 2.5 mmol) was dissolved in 100 mL ethylacetate. 10% Pd/C (500 mg) was added. The mixture was stirred under H₂balloon for overnight (20 h). It was filtered through celite. The celitewas washed thoroughly using ethyl acetate. The filtrate was concentratedand subjected to silica flash column to isolate compound 187.6 (650 mg,79%) as solid. MS found for C₁₆H₃₀N₂O₅ as (M+H)⁺ 331.3.

Step 7: Compound 187.6 (72 mg, 0.22 mmol) was dissolved in 10 mL dryDCM. To it were added 4 A molecular sieve (activated) (100 mg), NMO(solid, 39 mg, 0.33 mmol) and at last TPAP (8 mg, 0.022 mmol). Themixture was stirred at RT for 2 h. It was concentrated and directlyloaded on silica flash column to isolate compound 187.7 (77 mg, 100%) assolid. MS found for C₁₆H₂₈N₂O₅ as (M+H)⁺ 329.3.

Step 8: Ketone 187.7 (134 mg, 0.41 mmol) was dissolved in 10 mL DCM. Toit was added DAST (110 μl, 0.82 mmol). The mixture was stirred forovernight at RT. It was directly loaded onto silica flash column toisolate compound 187.8 (MS found for C₁₆H₂₈F₂N₂O₄ as (M+H)⁺ 351.2) using20% ethylacetate in hexane. It was then treated with neat TFA at RT for1 h and concentrated to dryness to afford compound 187.9. MS found forC₆H₁₂F₂N₂ as (M+H)⁺ 151.2.

Step 9: Compound 187.10 (72 mg, 0.2 mmol) was dissolved in 3 mL NMP. Toit was added DIEA (350 μL, 2 mmol) and all the crude compound 187.9 madein Step 8. The mixture was stirred at 90° C. for 2 h and subjected toreverse phase prep HPLC to isolate the title compound 187 (racemic). MSfound for C₁₈H₂₂F₂N₆O as (M+H)⁺ 377.2. UV 2=239, 292 nm. NMR (CD₃OD): δ8.51 (s, 1H), 7.45 (m, 1H), 7.34 (m, 1H), 7.25 (m, 1H), 7.02 (m, 1H),4.61 (m, 1H), 4.10 (m, 1H), 2.36 (s, 3H), 1.8-2.1 (m, 6H) ppm.

Example 3074-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,2R)-2-amino-3,3-difluorocyclohexylamino)pyrimidine-5-carboxamide(racemic)

The title compound was prepared using the same chemistry shown forExample 306. MS found for C₁₉H₂₁F₂N₉O as (M+H)⁺ 430.4. UV λ=250 nm.

Example 3082-((1R,2R,3R)-2-amino-3-fluorocyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic)

Step 1: Compound 187.6 (described in Example 306) (350 mg, 1.06 mmol)was dissolved in 20 mL DCM. At −78° C., to it was added a solution ofDAST (280 μL, 2.12 mmol) in 10 mL DCM dropwise. The mixture was stirredin the cold bath for 45 min, diluted with ethyl acetate, washed withbrine, concentrated and subjected by flash column to isolate compound189.1 in 30% yield. MS found for C₁₆H₂₉FN₂O₄ as (M+H)⁺ 333.2. It wastreated with 3 mL neat TFA for 20 min. The mixture was concentrated todryness to afford crude compound 189.2. MS found for C₆H₁₃FN₂ as (M+H)⁺133.1.

Step 2: Compound 187.10 (108 mg, 0.3 mmol) was dissolved in 3 mL NMP. Toit was added DIEA (260 μL, 1.5 mmol) and all the crude compound 189.2made in Step 1. The mixture was stirred at 90° C. for 2 h and subjectedto reverse phase prep HPLC to isolate the title compound 189 (racemic).MS found for C₁₈H₂₃FN₆O as (M+H)⁺ 359.2. UV 2=240, 292 nm. NMR (CD₃OD):δ 8.51 (s, 1H), 7.45 (m, 1H), 7.38 (m, 1H), 7.24 (m, 1H), 6.97 (m, 1H),4.83-4.78 (m, 2H), 3.65 (m, 1H), 2.36 (s, 3H), 1.8-2.2 (m, 6H) ppm.

Example 3094-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1R,2R,3R)-2-amino-3-fluorocyclohexylamino)pyrimidine-5-carboxamide(racemic)

The title compound was prepared using the same chemistry shown forExample 308. MS found for C₁₉H₂₂FN₉O as (M+H)⁺ 412.4. UV λ=251 nm.

Example 3102-((1R,2R,3S)-2-amino-3-hydroxycyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic, 191A) and2-((1S,2S,6R)-2-amino-6-hydroxycyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic, 191B)

Step 1: Compound 187.4 (described in Example 306) (180 mg, 0.60 mmol)was dissolved in 50 mL ethyl acetate. To it was added 10% Pd/C (100 mg)and the mixture was stirred for overnight under a hydrogen balloon. Themixture was filtered through celite, which was then thoroughly washedwith methanol. The filtrate was concentrated in vacuo to afford compound191.1 in quantitative yield as oil. MS found for C₁₂H₂₈N₂OSi as(M+H)⁺245.3.

Step 2: Compound 191.1 from Step 1 was dissolved in 5 mL NMP. To it wereadded DIEA (313 μL, 1.8 mmol) and compound 187.10 (108 mg, 0.3 mmol).The mixture was stirred at 90° C. for 90 min. To it were then added 10mL methanol, 10 mL TFA and 3 mL water. The mixture was stirred at 80° C.for 2 h. It was concentrated in vacuo and subjected to reverse prep HPLCto isolate racemic title compounds 191.A and 191.B. Ratio of 191.A (lesspolar) and 191.B was 7:1 by analytical HPLC. Compound 191.A: MS foundfor C₁₈H₂₄N₆O₂ as (M+H)⁺ 357.3. UV 2=243, 294 nm. Compound 191.B: MSfound for C₁₈H₂₄N₆O₂ as (M+H)⁺ 357.3. UV λ=243, 290 nm.

Example 3114-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1R,2R,3S)-2-amino-3-hydroxycyclohexylamino)pyrimidine-5-carboxamide(racemic, 192.A) and4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((1S,2S,6R)-2-amino-6-hydroxycyclohexylamino)pyrimidine-5-carboxamide(racemic, 192.B)

The two title racemic compounds were prepared using the same chemistryshown for Example 310. Ratio of 192.A (less polar) and 192.B was 1.6:1by analytical HPLC. Compound 192.A: MS found for C₁₉H₂₃N₉O₂ as (M+H)⁺410.4. UV λ=250 nm. Compound 192.B: MS found for C₁₉H₂₃N₉O₂ as (M+H)⁺410.4. UV λ=249 nm.

Example 3122-((1R,2R,3S)-2-amino-3-methoxycyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic, 193.A) and2-((1S,2S,6R)-2-amino-6-methoxycyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide(racemic, 193.B)

Step 1: Compound 187.4 (described in Example 306) (1.21 g, 4.1 mmol) wasdissolved in 60 mL dry THF. To it was added TBAF (1.0 M in THF, 8.2 mL,8.2 mmol). The mixture was stirred at RT for 2 h. It was concentratedand loaded onto silica flash column to isolate compound 193.1 (854 mg,99%) as solid.

Step 2: Compound 193.1 (124 mg, 0.68 mmol) was dissolved in 10 mL dryTHF. To it were added NaH (60 wt % in mineral oil, 55 mg, 1.36 mmol) and5 min later iodomethane (0.42 mL, 6.8 mmol). The mixture was stirred for1 h. It was concentrated in vacuo and loaded onto silica flash column toisolate compound 193.2 (115 mg, 85%) as oil.

Step 3: Compound 193.2 (115 mg, 0.58 mmol) made in Step 2 was dissolvedin 80 mL ethyl acetate. To it was added 100 mg 10% Pd/C and the mixturewas stirred under H₂ balloon for overnight. It was filtered throughcelite, and the celite was thoroughly washed with methanol. The filtratewas concentrated in vauo to give compound 193.3.

Step 4: All the compound 193.3 made in Step 3 was dissolved in 3 mL NMP.To it were added compound 187.10 (180 mg, 0.5 mmol) and DIEA (0.26 mL,1.5 mmol). The mixture was stirred at 90° C. for 1 h to give a mixtureof 193.A (less polar) and 193.B in ratio of 6:1. The two racemic titlecompounds were isolated using reverse phase prep HPLC. Compound 193.A:MS found for C₁₉H₂₆N₆O₂, as (M+H)⁺ 371.3. UV λ=247, 295 nm. Compound193.B: MS found for C₁₉H₂₆N₆O₂ as (M+H)⁺ 371.3. UV λ=242, 289 nm.

Example 3132-((1s,4s)-4-aminocyclohexylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

To a mixture of trans-4-aminocyclohexanol (2.07 g, 13.6 mmol) and NaHCO₃(3.50 g, 41.7 mmol) in H₂O (20 mL) at room temperature, a solution ofbenzyl chloroformate (1.92 mL, 13.6 mmol) in dioxane (15 mL) was added.The mixture was stirred at room temperature for 20 h. The whiteprecipitate was collected as benzyl (1R,4R)-4-hydroxycyclohexylcarbamate(3.37 g).

To a suspension of benzyl (1R,4R)-4-hydroxycyclohexylcarbamate (1.14 g,4.58 mmol) and triethylamine (1.30 mL, 9.34 mmol) in CH₂Cl₂ (15 mL) atroom temperature, methanesulfonyl chloride (0.425 mL, 5.49 mmol) wasadded. The mixture was stirred at room temperature for 20 h. Moremethanesulfonyl chloride (0.425 mL, 5.49 mmol) and triethylamine (1.00mL) were added. Stirring was continued for 48 h. The reaction solutionwas washed with 5% NaHCO₃, then with 1 N HCl. The organic phase wasseparated, dried over Na₂SO₄, concentrated in vacuo to give(1R,4R)-4-(benzyloxycarbonyl)cyclohexyl methanesulfonate as a solid(1.13 g).

A mixture of (1R,4R)-4-(benzyloxycarbonyl)cyclohexyl methanesulfonate(1.13 g, 3.46 mmol) and NaN3 (0.674 g, 10.4 mmol) in DMF (10 mL) wasstirred at 100 C for 20 h. Water and EtOAc were added. The organic phasewas separated, washed with water, dried over Na₂SO₄, concentrated invacuo to give benzyl (1s,4s)-4-azidocyclohexylcarbamate (0.819 g).

A mixture of benzyl (1s,4s)-4-azidocyclohexylcarbamate (400 mg, 1.46mmol) and Pd—C (10%, 70 mg) in MeOH (20 mL) (containing 5 drops of 6NHCl) was hydrogenated under balloon hydrogen for 20 h. It was filteredthrough celite. The filtrate was concentrated in vacuo to give(1s,4s)-cyclohexane-1,4-diamine (223 mg).

To a solution of (1s,4s)-cyclohexane-1,4-diamine (60 mg, 0.40 mmol) andTEA (0.208 mL, 1.50 mmol) in DMF (2 mL) and MeOH (1 mL), compound2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(m-tolylamino)pyrimidine-5-carboxamide(65 mg, 0.18 mmol) was added. The mixture was stirred at roomtemperature for 20 h. It was concentrated in vacuo. The residue waspurified by HPLC to give the titled compound (30 mg). MS 341.2 (M+H); UV247.8, 298.8.

Example 3142-((4-aminotetrahydro-2H-pyran-4-yl)methylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

Compound 187.10 (as shown in Example 306) (70 mg, 0.19 mmol) wasdissolved in 3 mL NMP. To it were added commercially available4-(aminomethyl)tetrahydro-2H-pyran-4-amine (78 mg, 0.38 mmol) and DIEA(0.20 mL, 1.14 mmol). The mixture was stirred at 90° C. for 2 h. Fromthis mixture the title compound was isolated using reverse phase prepHPLC. MS found for C₁₈H₂₄N₆O₂ as (M+H)⁺ 357.3. UV λ=240 nm.

Example 315(R)-4-(1H-indazol-6-ylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

Step 1: To a stirring solution of carboxylic acid 165.1 (85 g, 540 mmol)in thionyl chloride (425 mL) was added pyridine (8.5 mL, 0.11 mmol),slowly. The reaction was stirred at 75° C. overnight at which time itwas concentrated and dried under vacuum to a light yellow powder. Thisyellow solid was slowly diluted with 750 mL of ethanol and refluxedovernight. The next day the reaction was determined to be complete byHPLC and then cooled in an ice bath and the solid filtered and washedwith diethyl ether affording ethyl ester 165.2 as an off-white powder(91 g, 87% for two steps). MS found for C₇H₈N₂O₄ as (M+H)⁺ 185.0.

Step 2: Ester 165.2 (22 g, 120 mmol) was dissolved in phosphorousoxychloride (60 mL, 600 mmol) and the mixture treated withN,N-diethylaniline (27 mL, 167 mmol) and the mixture heated to 105° C.until the reaction was determined to be complete by HPLC. It was thencooled to RT and slowly added to 1 L of crushed ice resulting in theformation of a beige precipitate which was collected by filtration anddried under vacuum affording dichloride 165.3 as a light yellow powder(22.5 g, 85%). ¹H NMR (DMSO-thud 6, 400 MHz): δ 9.13 (s, 1H), 4.37 (q,2H), 1.32 (t, 3H).

Step 3: Dichloropyrimidine 165.3 (1.04 g, 4.7 mmol) was dissolved in NMP(30 mL) and stirred in ice bath. To it were added 6-aminoindazole 165.4(690 mg, 5.2 mmol) and then dropwise ethyldiisopropylamine (DIEA, 1.64mL, 9.4 mmol). The mixture was stirred for 40 minutes, and to it wasadded sodium thiomethoxide (660 mg, 9.4 mmol). The mixture was stirredfor overnight, diluted with ethyl acetate, washed with brine threetimes, and concentrated in vacuo to give crude compound 165.5 as a lightbrown solid in quantitative yield. MS found for C₁₅H₁₅N₅O₂S as (M+H)⁺330.1.

Step 4: Ethyl ester 165.5 (4.7 mmol) was dissolved in 60 mL THF. To itwere added lithium hydroxide hydrate (236 mg, 5.6 mmol) and 20 mL water.The mixture was stirred for overnight and to it was carefully added 1NHCl solution till pH reaching 2. The mixture was concentrated in vacuoto remove THF. White solid crashed out and was isolated using a Büchnerfunnel. It was washed with water and dried in vacuum oven to givecompound 165.6 (1.14 g, 81%) as a white solid. MS found for C₁₃H₁₁N₅O₂Sas (M+H)⁺ 302.1.

Step 5: Carboxylic acid 165.6 (1.14 g, 3.8 mmol) was dissolved in 30 mLDMF. To it were added EDC hydrochloride (1.09 g, 5.7 mmol) and HOBthydrate (770 mg, 5.7 mmol). The mixture was stirred at RT for 1 hour. Toit was then added ammonia (commercial 0.5N solution in dioxane, 22 mL,11.4 mmol). The mixture was stirred for 2 hours. It was thenconcentrated in vacuo and taken into water and ethyl acetate. Theorganic phase was separated and washed with brine four times. Theorganic phase was then dried over MgSO₄ and concentrated in vacuo toafford compound 165.7 as a light yellow solid (820 mg, 72%). MS foundfor C₁₃H₁₂N₆OS as (M+H)⁺ 301.1.

Step 6: Compound 165.7 (40 mg, 0.13 mmol) was dissolved in 3 mL NMP. Toit was added MCPBA (65% pure, 53 mg, 0.18 mmol). It was stirred at RTfor 45 minutes. To it then were added H-D-Leu-NH₂HCl (108 mg, 0.65 mmol)and DIEA (230 μL, 1.3 mmol). The mixture was stirred for 90 minutes at120° C. bath. This mixture was then subjected to preparative HPLC toisolate the title compound 165. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.2.UV λ=246, 303 nm.

Example 316(R)-4-(1H-indazol-6-ylamino)-2-(1-amino-1-oxopropan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₅H₁₆N₈O₂ as (M+H)⁺ 341.2. UV 2=245, 302nm.

Example 317(R)-4-(1H-indazol-6-ylamino)-2-(2-amino-2-oxo-1-phenylethylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Phe-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₂₀H₁₈N₈O₂ as (M+H)⁺ 403.2. UV 2=249, 298nm.

Example 318(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(quinolin-6-ylamino)pyrimidine-5-carboxamide

Step 1: Dichloropyrimidine 169.1 (500 mg, 2.3 mmol) was dissolved in NMP(20 mL) and stirred in ice bath. To it were added 6-aminoquinoline 169.2(390 mg, 2.7 mmol) and then dropwise ethyldiisopropylamine (DIEA, 0.72mL, 4.1 mmol). The mixture was stirred for 2 hours, diluted with ethylacetate, washed with brine three times, and concentrated in vacuo togive crude compound 169.3 as a light brown solid in quantitative yield.MS found for C₁₆H₁₃ClN₄O₂ as (M+H)⁺ 329.1.

Step 2: Ethyl ester 169.3 (2.3 mmol) was dissolved in 30 mL THF. To itwere added lithium hydroxide hydrate (193 mg, 4.6 mmol) and 6 mL water.The mixture was stirred for 7 hours and to it was carefully added 1N HClsolution till pH reaching 5. The mixture was concentrated in vacuo toremove THF and was extracted with ethyl acetate 5 times. The organicphases were combined, dried and concentrated in vacuo to give crude acid169.4. MS found for C₁₄H₉ClN₄O₂ as (M+H)⁺ 301.1.

Step 3: Carboxylic acid 169.4 (220 mg, 0.73 mmol) was dissolved in 18 mLNMP. To it were added EDC hydrochloride (210 mg, 1.1 mmol) and HOBthydrate (150 mg, 1.1 mmol). The mixture was stirred at RT for 1 hour. Toit was then added ammonia (commercial 0.5N solution in dioxane, 7.3 mL,3.65 mmol). The mixture was stirred for 2.5 hours. It was thenconcentrated in vacuo and taken into water and ethyl acetate. Theorganic phase was separated and washed with brine three times. Theorganic phase was then dried over MgSO₄ and concentrated in vacuo toafford compound 169.5 as a solid (180 mg, 62%). MS found for C₂₀H₁₄N₈O₂as (M+H)⁺ 399.1.

Step 6: Compound 169.5 (71 mg, 0.18 mmol) was dissolved in 3 mL NMP. Toit were added H-D-Leu-NH₂HCl (150 mg, 0.90 mmol) and DIEA (0.31 mL, 1.8mmol). The mixture was stirred for 90 minutes at 120° C. bath. Thismixture was then subjected to preparative HPLC to isolate the racemictitle compound 169. MS found for C₂₀H₂₃N₇O₂ as (M+H)⁺ 394.2. UV λ=241,283 nm.

Example 319(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 6-aminobenzothiophene to replace6-aminoindazole 165.4. MS found for C₁₈H₂₁N₇O₂S as (M+H)⁺ 400.2. UVλ=243, 303 nm.

Example 320(R)-4-(1H-indol-5-ylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 6-aminoindole to replace6-aminoindazole 165.4. MS found for C₁₉H₂₃N₇O₂ as (M+H)⁺ 382.2. UV λ=245nm.

Example 321(R)-4-(1H-indazol-5-ylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 5-aminoindazole to replace6-aminoindazole 165.4. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.2. UVλ=247, 301 nm.

Example 322(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-(methylsulfonyl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic scheme inExample 315 with 3-(methylsulfonyl)aniline to replace 6-aminoquinoline169.2. MS found for C₁₈H₂₄N₆O₄S as (M+H)⁺ 421.2. UV λ=249, 285 nm.

Example 323(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(methylsulfonyl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(methylsulfonyl)aniline to replace6-aminoquinoline 169.2. MS found for C₁₈H₂₄N₆O₄S as (M+H)⁺ 421.2. UVλ=250, 300 nm.

Example 324(R)-4-(4-(4-acetylpiperazin-1-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with1-(4-(4-aminophenyl)piperazin-1-yl)ethanone to replace 6-aminoquinoline169.2. MS found for C₂₃H₃₂N₈O₃ as (M+H)⁺ 469.3. UV λ=243, 296 nm.

Example 325((R)-4-(1H-benzo[d]imidazol-6-ylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 1H-benzo[d]imidazol-6-amine to replace6-aminoquinoline 169.2. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.2. UVλ=244, 297 nm.

Example 327(R)-4-(1H-indazol-5-ylamino)-2-(1-amino-1-oxopropan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated Example 315 with H-D-Ala-NH₂HCl to replace H-D-Leu-NH₂HCl.MS found for C₁₅H₁₆N₈O₂ as (M+H)⁺ 341.2. UV λ=245, 299 nm.

Example 328(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(N-methylacetamido)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with N-(4-aminophenyl)-N-methylacetamide toreplace 6-aminoquinoline

169.2. MS found for C₂₀H₂₇N₇O₃ as (M+H)⁺ 414.2. UV λ=245, 296 nm.

Example 329(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(4-(N-methylacetamido)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₇H₂₁N₇O₃ as (M+H)⁺ 372.2. UV λ=244, 295nm.

Example 330(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₅H₁₅N₇O₂S as (M+H)⁺ 358.1. UV λ=244, 303nm. NMR (CD₃OD): δ 9.22 (s, 1H), 8.51 (m, 2H), 8.07 (d, J=8.8 Hz, 1H),7.71 (d, J=8.8 Hz, 1H), 4.44 (m, 1H), 1.55 (d, J=6.8 Hz, 3H) ppm.

Example 331(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 5-aminobenzothiophene to replace6-aminobenzothiophene. MS found for C₁₅H₁₅N₇O₂S as (M+H)⁺ 358.1. UVλ=249, 292 nm. NMR (CD₃OD): δ 9.34 (s, 1H), 8.67 (s, 1H), 8.51 (s, 1H),8.10 (d, J=8.8 Hz, 1H), 7.59 (d, J=8.8 Hz, 1H), 4.64 (m, 1H), 1.55 (d,J=7.2 Hz, 3H) ppm.

Example 332(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 5-aminobenzothiophene to replace6-aminobenzothiophene. MS found for C₁₈H₂₁N₇O₂S as (M+H)⁺ 400.2. UVλ=247, 295 nm.

Example 333(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(imidazo[1,2-a]pyridin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with imidazo[1,2-a]pyridin-6-amine toreplace 6-aminobenzothiophene. MS found for C₁₅H₁₆N₈O₂ as (M+H)⁺ 341.2.UV λ=250 nm.

Example 334(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(imidazo[1,2-a]pyridin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with imidazo[1,2-a]pyridin-6-amine toreplace 6-aminoindazole 165.4. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.2.UV 2=252 nm.

Example 335(R)-4-(1H-indol-5-ylamino)-2-(1-amino-1-oxopropan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₆H₁₇N₇O₂ as (M+H)⁺ 340.2. UV λ=224 nm.NMR (CD₃OD): δ 8.38 (s, 1H), 7.84 (m, 1H), 7.40 (m, 1H), 7.26 (m, 2H),6.51 (s, 1H), 4.44 (m, 1H), 1.50 (d, J=7.6 Hz, 3H) ppm. 8.82 (s, 1H),8.74 (m, 1H), 8.61 (s, 1H), 8.10 (d, J=8.8 Hz, 1H), 7.87 (d, J=8.4 Hz,1H), 4.54 (m, 1H), 3.82 (m, 1H), 1.99-1.62 (m, 8H) ppm.

Example 336(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(2,3-dihydrobenzo[b][1,4]dioxin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 2,3-dihydrobenzo[b][1,4]dioxin-6-amineto replace 6-aminoquinoline 169.2. MS found for C₁₉H₂₄N₆O₄ as (M+H)⁺401.2. UV λ=243, 310 nm.

Example 337(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(2,3-dihydrobenzo[b][1,4]dioxin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₆H₁₈N₆O₄ as (M+H)⁺ 359.2. UV λ=243, 312nm.

Example 338(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(quinoxalin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with quinoxalin-6-amine to replace6-aminoindazole 165.4. MS found for C₁₉H₂₂N₈O₂ as (M+H)⁺ 395.2. UV λ=243nm.

Example 339(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(quinoxalin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with H-D-Ala-NH₂HCl to replaceH-D-Leu-NH₂HCl. MS found for C₁₆H₁₆N₈O₂ as (M+H)⁺ 353.2. UV λ=242 nm.

Example 340(R)-4-(4-(1H-pyrazol-1-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(1H-pyrazol-1-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₀H₂₄N₈O₂ as (M+H)⁺ 409.2. UVλ=241, 308 nm.

Example 341(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(thiazol-4-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₀H₂₃N₇O₂S as (M+H)⁺ 426.2. UVλ=240, 314 nm.

Example 342(R)-4-(4-(1,2,3-thiadiazol-4-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(1,2,3-thiadiazol-4-yl)aniline toreplace 6-aminoindazole 165.4. MS found for C₁₉H₂₂N₈O₂S as (M+H)⁺ 427.2.UV λ=233, 311 nm.

Example 343(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(benzo[c][1,2,5]thiadiazol-5-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with benzo[c][1,2,5]thiadiazol-5-amine toreplace 6-aminoquinoline 169.2. MS found for C₁₇H₂₀N₈O₂S as (M+H)⁺400.2. UV λ=240 nm.

Example 344(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(pyridin-2-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(pyridin-2-yl)aniline to replace6-aminoquinoline 169.2. MS found for C₂₂H₂₅N₇O₂ as (M+H)⁺ 420.2. UV2=244, 314 nm.

Example 345(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-morpholinophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-morpholinoaniline to replace6-aminoquinoline 169.2. MS found for C₂₁H₂₉N₇O₃ as (M+H)⁺ 428.2. UVλ=244, 294 nm.

Example 346(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-fluoro-4-morpholinophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-fluoro-4-morpholinoaniline to replace6-aminoquinoline 169.2. MS found for C₂₁H₂₈FN₇O₃ as (M+H)⁺ 446.2. UVλ=240, 310 nm.

Example 347(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-morpholinophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-morpholinoaniline to replace6-aminoquinoline 169.2. MS found for C₂₁H₂₉N₇O₃ as (M+H)⁺ 428.2. UVλ=246 nm.

Example 348a(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(piperidin-1-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(piperidin-1-yl)aniline to replace6-aminoquinoline 169.2. MS found for C₂₂H₃₁N₇O₂ as (M+H)⁺ 426.2. UVλ=247, 291 nm.

Example 348b(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(pyrrolidin-1-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(pyrrolidin-1-yl)aniline to replace6-aminoquinoline 169.2. MS found for C₂₁H₂₉N₇O₂ as (M+H)⁺ 412.2. UVλ=247, 292 nm.

Example 349a(R)-4-(4-(1H-imidazol-1-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(1H-imidazol-1-yl)aniline to replace6-aminoquinoline 169.2. MS found for C₂₀H₂₄N₈O₂ as (M+H)⁺ 409.2. UVλ=247, 297 nm.

Example 349b(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-(pyrimidin-5-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-(pyrimidin-5-yl)aniline to replace6-aminoquinoline 169.2. MS found for C₂₁H₂₄N₈O₂ as (M+H)⁺ 421.2. UVλ=247 nm.

Example 350(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-(pyridin-4-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-(pyridin-4-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₂H₂₅N₇O₂ as (M+H)⁺ 420.2. UVλ=241, 314 nm.

Example 351(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-(pyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-(pyridin-3-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₂H₂₅N₇O₂ as (M+H)⁺ 420.2. UV λ=250nm.

Example 352(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(3-(pyridin-3-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-(1H-imidazol-1-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₀H₂₄N₈O₂ as (M+H)⁺ 409.2. UV2=243, 284 nm.

Example 353(R)-4-(3-(1′-1-pyrazol-1-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 3-(1H-pyrazol-1-yl)aniline to replace6-aminoindazole 165.4. MS found for C₂₀H₂₄N₈O₂ as (M+H)⁺ 409.2. UV λ=250nm.

Example 354(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(4-(oxazol-5-yl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(oxazol-5-yl)aniline to replace6-aminobenzothiophene. MS found for C₁₇H₁₇N₇O₃ as (M+H)⁺ 368.2.

Example 355(R)-4-(4-(1H-1,2,4-triazol-1-yl)phenylamino)-2-(1-amino-1-oxopropan-2-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 4-(1H-1,2,4-triazol-1-yl)aniline toreplace 6-aminobenzothiophene. MS found for C₁₆H₁₇N₉O₂ as (M+H)⁺ 368.2.

Example 356(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(2-(morpholinomethyl)quinolin-6-ylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 315 with 2-(morpholinomethyl)quinolin-6-amine toreplace 6-aminobenzothiophene. MS found for C₂₂H₂₆N₈O₃ as (M+H)⁺ 451.2.

Example 357(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)pyrimidine-5-carboxamide

Compound 179.6 (100 mg, 0.31 mmol) was dissolved in 5 mL NMP. To it wasadded MCPBA (97 mg, 0.37 mmol) and the mixture was stirred for 30 min.To it was added DIEA (0.32 mL, 1.86 mmol) and then commerciallyavailable H-D-Leu-NH₂.HCl (155 mg, 0.93 mmol). The mixture was stirredat 120° C. for 3 h. From this mixture the title compound was isolatedusing reverse phase prep HPLC. MS found for C₁₉H₂₃N₉O₂ as (M+H)⁺ 410.3.UV λ=251 nm.

Example 358(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-1-oxobutan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 357. MS found for C₁₇H₁₉N₉O₂ as (M+H)⁺ 382.3. UV λ=250 nm.

Example 359(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-1-oxopropan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 357. MS found for C₁₆H₁₇N₉O₂ as (M+H)⁺ 368.3. UV λ=251 nm.

Example 3604-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(2-amino-2-oxoethylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 357. MS found for C₁₅H₁₅N₉O₂ as (M+H)⁺ 354.3. UV λ=250 nm.

Example 361(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(3,6-dihydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry shown forExample 357. MS found for C₂₂H₂₈N₆O₃ as (M+H)⁺ 425.4. UV 2=240, 313 nm.

Example 362(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(tetrahydro-2H-pyran-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared from Example 361 using standardcatalysis hydrogenation by treating the solution of Example 261 inmethanol with 10% Pd/C under H₂ balloon for overnight. MS found forC₂₂H₃₀N₆O₃ as (M+H)⁺ 427.4. UV λ=244, 298 nm.

Example 363(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(tetrahydro-2H-pyran-4-yloxy)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 315 with the corresponding aniline which wascommercially available. MS found for C₂₂H₃₀N₆O₄ as (M+H)⁺ 443.4. UVλ=244, 295 nm.

Example 364(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(tetrahydro-2H-pyran-4-ylsulfonyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 315 with the corresponding aniline (synthesisshown in for compound 266). MS found for C₂₂H₃₀N₆O₅S as (M+H)⁺ 491.4. UVλ=251, 302 nm.

Example 365(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-(morpholinosulfonyl)phenylamino)pyrimidine-5-carboxamide

Example 366(R)-2-(1-amino-3-methyl-1-oxobutan-2-ylamino)-4-(cyclobutylamino)pyrimidine-5-carboxamide

Benzotriazolyl ether 1.7 (509 mg, 0.17 mmol), (R)-valine hydrochloride(30 mg, 0.26 mmol), diisopropylethylamine (89 uL, 0.51 mmol) and 5 mL of1,4-dioxane were combined and heated to 120° C. overnight. The reactionmixture was then cooled, diluted with water and purified by preparativeHPLC to give the desired compound. MS found for C₁₄H₂₂N₆O₂ as (M+H)⁺307.3.

Example 367(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(4-methylthiophen-2-ylamino)pyrimidine-5-carboxamide

The titled compound was then synthesized analogously by using4-methylthiophen-2-amine. MS 363.3 (M+H); UV 201.6, 244.3, 326.1 nm.

Example 368(R)-2-(1-amino-3-methyl-1-oxobutan-2-ylamino)-4-(4-methylthiophen-2-ylamino)pyrimidine-5-carboxamide

The titled compound was then synthesized analogously by using4-methylthiophen-2-amine. MS 349.3 (M+H); UV 202.9, 251.0, 324.8 nm.

Example 369(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 318. MS found for C₂₀H₂₅N₇O₂ as (M+H)⁺ 396.3.UV: λ=216.9, 244.0.

Example 370(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 318. MS found for C₁₉H₂₄N₈O₂ as (M+H)⁺ 397.4.UV: λ=213.3, 244.0, 333.1.

Example 371(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 318. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.4.UV: λ=241.6, 318.8.

Example 372(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 318. MS found for C₁₉H₂₄N₈O₂ as (M+H)⁺ 397.4.UV: λ=207.5, 244.0, 325.9.

Example 373(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

Step 1 & 2 are same as described for Example 100.

Step 3: Dichloropyrimidine 165.3 (5.09 g, 22.6 mmol) was dissolved inacetonitrile (50 mL) and stirred in ice bath. To it were added3,5-dimethylaniline (2.19 g, 18.2 mmol) and then dropwiseethyldiisopropylamine (DIEA, 5.74 mL, 33.9 mmol). The mixture wasstirred at room temperature overnight, diluted with ethyl acetate,washed with brine three times, and concentrated in vacuo to give crudecompound 4 as a light brown solid 2.4 gm (88% yield). MS found forC₁₅H₁₆ClN₃O₂ as (M+H)⁺ 306.09.

Step 4: Ethyl ester 4 (9.8 mmol) was dissolved in 20 mL THF. To it wereadded lithium hydroxide hydrate (530 mg, 10.2 mmol) and 15 mL water. Themixture was stirred for overnight and to it was carefully added 1N HClsolution till pH reaching 2. The mixture was concentrated in vacuo toremove THF. White solid crashed out and was isolated using a Büchnerfunnel. It was washed with water and dried in vacuum oven to givecompound 5 (2.14 g, 81%) as a white solid. MS found for C₁₃H₁₂ClN₃O₂ as(M+H)⁺ 278.1.

Step 5: Carboxylic acid 5 (1.09 g, 3.6 mmol) was dissolved in 30 mL DMF.To it were added EDC hydrochloride (1.24 g, 6.4 mmol) and HOBt hydrate(870 mg, 6.4 mmol). The mixture was stirred at RT for 1 hour. Thereaction mixture was cooled to 0° C. to it was then added ammonia(commercial 0.4M solution in dioxane, 16 mL, 8.3 mmol). The mixture wasstirred for 2 hours. It was then concentrated in vacuo and taken intowater and ethyl acetate. The organic phase was separated and washed withbrine four times. The organic phase was then dried over MgSO₄ andconcentrated in vacuo to afford compound 6 as a solid (150 mg, 55%). MSfound for C₁₉H₁₇N₇O₂ as (M+H)⁺ 376.1.

Step 6: Compound 6 (188 mg, 0.5 mmol) was dissolved in 3 mL NMP. To itwere added H-D-Val-NH₂HCl (174 mg, 1.5 mmol) and DIEA (0.200 mL, 1.0mmol). The mixture was stirred for 90 minutes at 120° C. bath. Thismixture was then subjected to preparative HPLC to isolate the titlecompound 7. MS found for C₁₈H₂₄N₆O₂ as (M+H)⁺ 357.20.

Example 374(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3,5-dimethoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3,5-dimethoxyaniline to replace3,5-dimethylaniline. MS found for C₁₈H₂₄N₆O₄ as (M+H)⁺ 388.

Example 375(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-bromophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-bromoaniline to replace3,5-dimethylaniline. MS found for C₁₆H₁₉BrN₆O₂ as (M+H)⁺ 407.

Example 376(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-ethoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-ethoxyaniline to replace3,5-dimethylaniline. MS found for C₁₈H₂₄N₆O₃ as (M+H)⁺ 373

Example 377(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-ethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-ethylaniline to replace3,5-dimethylaniline. MS found for C₁₈H₂₄N₆O₂ as (M+H)⁺ 356.3

Example 378(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-(trifluoromethyl)phenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-trifluoromethylaniline to replace3,5-dimethylaniline. MS found for C₁₇H₁₉F₃N₆O₂ as (M+H)⁺ 396.3

Example 379 (R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-cyanophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-aminobenzonitrile to replace3,5-dimethylaniline. MS found for C₁₇H₁₉N₇O₂ as (M+H)⁺ 353.38

Example 380 (R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-methoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with 3-methoxyaniline to replace3,5-dimethylaniline. MS found for C₁₇H₂₂N₆O₃ as (M+H)⁺ 358.4

Example 381(R)-2-(-amino-3-methyl-1oxobutan-2-ylamino)-4-(3-(m-tolylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with m-toluidine to replace3,5-dimethylaniline. MS found for C₁₇H₂₂N₆O₂ as (M+H)⁺ 343

Example 382(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ala-NH₂HCl to replaceH-D-Val-NH₂HCl. MS found for C₁₆H₂₀N₆O₂ as (M+H)⁺ 328.2.

Example 383(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(3,5-dimethoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 382 with 3,5-dimethoxyaniline to replace3,5-dimethylaniline. MS found for C₁₆H₂₀N₆O₄ as (M+H)⁺ 360.3.

Example 384(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(3-bromophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 382 with 3-bromoaniline to replace3,5-dimethylaniline. MS found for C₁₄H₁₅BrN₆O₄ as (M+H)⁺ 360.3.

Example 385(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(3-ethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 382 with 3-ethylaniline to replace3,5-dimethylaniline. MS found for C₁₆H₂₀N₆O₂ as (M+H)⁺ 328.3.

Example 386(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 382 with m-toluidine to replace3,5-dimethylaniline. MS found for C₁₅H₁₈N₆O₂ as (M+H)⁺ 314.34.

Example 387(R)-2-(1-amino-1-oxopropan-2-ylamino)-4-(3-methoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 382 with 3-methoxyaniline to replace3,5-dimethylaniline. MS found for C₁₅H₁₈N₆O₃ as (M+H)⁺ 330.34.

Example 388(R)-2-(2-amino-2-oxo-1-phenylethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Phe-NH₂HCl to replaceH-D-Val-NH₂HCl. MS found for C₂₁H₂₂N₆O₂ as (M+H)⁺ 328.2.

Example 389(R)-2-(2-amino-2-oxo-1-phenylethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Phe-NH₂HCl to replaceH-D-Val-NH₂HCl and 3,5-dimethoxyaniline to replace 3,5-dimethylaniline.MS found for C₂₁H₂₂N₆O₄ as (M+H)⁺ 423.

Example 390(R)-2-(2-amino-2-oxo-1-phenylethylamino)-4-(3-methoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Phe-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-methoxyaniline to replace 3,5-dimethylaniline. MSfound for C₂₀H₂₀N₆O₃ as (M+H)⁺ 392.4.

Example 391(R)-2-(2-amino-2-oxo-1-phenylethylamino)-4-(3-trifluoromethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Phe-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-trifluoromethylaniline to replace3,5-dimethylaniline. MS found for C₂₀H₁₇F₃N₆O₂ as (M+H)⁺430.39.

Example 392(R)-2-(2-amino-2-oxo-1-phenylethylamino)-4-(3-bromophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Phe-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-bromoaniline to replace 3,5-dimethylaniline. MSfound for C₁₋₁₉H₁₇BrN₆O₂ as (M+H)⁺ 441.29.

Example 393(R)-2-(1-amino-3-hydroxy-1-oxo-2-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ser-NH₂HCl to replaceH-D-Val-NH₂HCl. MS found for C₁₆H₂₀N₆O₃ as (M+H)⁺ 344.37.

Example 394(R)-2-(1-amino-3-hydroxy-1-oxo-2-ylamino)-4-(3-ethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ser-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-ethylaniline to replace 3,5-dimethylaniline. MSfound for C₁₆H₂₀N₆O₃ as (M+H)⁺ 344.37.

Example 396(R)-2-(1-amino-3-hydroxy-1-oxo-2-ylamino)-4-(3-m-tolylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ser-NH₂HCl to replaceH-D-Val-NH₂HCl and m-toluidine to replace 3,5-dimethylaniline. MS foundfor C₁₅H₁₈N₆O₃ as (M+H)⁺ 330.34.

Example 397(R)-2-(1-amino-3-hydroxy-1-oxo-2-ylamino)-4-(3-bromophenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ser-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-bromoaniline to replace 3,5-dimethylaniline. MSfound for C₁₄H₁₅BrN₆O₃ as (M+H)⁺ 395.22.

Example 398(R)-2-(1-amino-3-hydroxy-1-oxo-2-ylamino)-4-(3-methoxyphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with H-D-Ser-NH₂HCl to replaceH-D-Val-NH₂HCl and 3-methoxyaniline to replace 3,5-dimethylaniline. MSfound for C₁₅H₁₈N₆O₄ as (M+H)⁺ 346.34.

Example 399(R)-2-(2-amino-2-oxo-1-m-tolylethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with (S)-2-amino-2-m-tolylacetamide toreplace H-D-Val-NH₂HCl. MS found for C₂₂H₂₄N₆O₂ as (M+H)⁺ 404.47.

Example 400(R)-2-(2-amino-1-(3-chlorophenyl)-2-oxoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with (S)-2-amino-2-(3-chlorophenyl)acetamideto replace H-D-Val-NH₂ HCl. MS found for C₂₁H₂₁ClN₆O₂ as (M+H)⁺ 424.89.

Example 401(R)-2-(2-amino-1-(3-fluorophenyl)-2-oxoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with (S)-2-amino-2-(3-fluorophenyl)acetamideto replace H-D-Val-NH₂ HCl. MS found for C₂₁H₂₁FN₆O₂ as (M+H)⁺ 408.43.

Example 402(R)-2-(2-amino-2-oxo-1-p-tolylethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with (S)-2-amino-p-tolylacetamide to replaceH-D-Val-NH₂HCl. MS found for C₂₂H₂₄N₆O₂ as (M+H)⁺ 404.47.

Example 403(R)-2-(2-amino-2-oxo-3(trifluoromethylphenyl)ethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with(S)-2-amino-2-(3-fluoromethylphenyl)acetamide to replace H-D-Val-NH₂HCl.MS found for C₂₂H₂₁F₃N₆O₂ as (M+H)⁺ 404.47.

Example 404(R)-2-(2-amino-2-oxo-1(pyridine-3-yl)ethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with (S)-2-amino-2-(pyridine-3-yl)acetamideto replace H-D-Val-NH₂HCl. MS found for C₂₀H₂₁N₇O₂ as (M+H)⁺ 391.43.

Example 405(R)-2-(2-amino-1(4-methoxyphenyl)-2-oxoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with(S)-2-amino-2-(4-methoxyphenyl)acetamide to replace H-D-Val-NH₂ HCl. MSfound for C₂₂H₂₄N₆O₃ as (M+H)⁺ 420.47.

Example 406(R)-2-(2-amino-1(3-hydroxyphenyl)-2-oxoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with(S)-2-amino-2-(3-hydroxyphenyl)acetamide to replace H-D-Val-NH₂ HCl. MSfound for C₂₁H₂₂N₆O₃ as (M+H)⁺ 406.44.

Example 407(R)-2-(2-amino-1(34-hydroxyphenyl)-2-oxoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The above compound was prepared using the same synthetic schemedemonstrated in Example 373 with(S)-2-amino-2-(4-hydroxyphenyl)acetamide to replace H-D-Val-NH₂ HCl. MSfound for C₂₁H₂₂N₆O₃ as (M+H)⁺ 406.44.

Example 408(R)-2-(1-amino-3-methyl-1-oxobutan-2-ylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

To a stirring solution of 368.1 (110. mg, 0.347 mmol) was added mCPBA(65%, 130 mg, 0.49 mmol). The reaction was stirred for 30 min at RT uponwhich time DIEA (˜0.3 mL) and commercially available 368.2 (180 mg, 1.18mmol) were added. The reaction was heated for 3 h at 120° C. in a sealedtube. The reaction was cooled, turned slightly acidic with TFA in water,and subjected to reverse phase preparative HPLC to yield the titlecompound (28 mg). MS found for C₁₇H₁₉N₇O₂S as (M+H)⁺ 386.3. UV 2=248,294 nm

Example 409(R)-2-(1-amino-1-oxobutan-2-ylamino)-4-(quinolin-6-ylamino)pyrimidine-5-carboxamide

Intermediate 369.1 was reacted with 346.1 according to Example 408.Reverse phase preparative HPLC afforded the title compound. MS found forC₁₈H₁₉N₇O₂ as (M+H)⁺ 366.2. UV λ=240, 283 nm. δ 1.15 (t, 3H), 1.90-2.10(m, 2H), 3.10-3.35 (m, 2H), 4.32-4.40 (m, 1H), 7.80-7.90 (m, 1H),8.07-8.18 (m, 2H), 8.59 (s, 1H), 8.70-8.75 (m, 1H), 8.95-9.05 (m, 2H).

Example 410(R)-2-(1-amino-3-methyl-1-oxobutan-2-ylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

Intermediate 371.1 was reacted with 368.2 according to the chemistrydescribed in Example 408. Reverse phase preparative HPLC afforded thetitle compound. MS found for C₁₇H₁₉N₇O₂S as (M+H)⁺ 386.4. UV λ=243, 302nm.

Example 411(R)-2-(1-amino-1-oxobutan-2-ylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

Intermediate 368.1 was reacted with 346.1 according to the chemistrydescribed in Example 408. Reverse phase preparative HPLC afforded thetitle compound. MS found for C₁₆H₁₇N₇O₂S as (M+H)⁺ 372.3. UV λ=246, 293nm

Example 412(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 417. MS found for C₂₀H₂₅N₇O₂ as (M+H)⁺ 396.3.UV: λ=216.9, 244.0.

Example 413(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 315. MS found for C₁₋₁₉H₂₄N₈O₂ as (M+H)⁺ 397.4.UV: λ=213.3, 244.0, 333.1.

Example 414(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 315. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.4.UV: λ=241.6, 318.8.

Example 415(R)-2-(1-amino-4-methyl-1-oxopentan-2-ylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 315. MS found for C₁₉H₂₄N₈O₂ as (M+H)⁺ 397.4.UV: λ=207.5, 244.0, 325.9.

Example 4162-(2-aminoethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

Compound 195.1 (64 mg, 0.17 mmol) was dissolved in 3 mL NMP. To it wasadded commercially available ethylenediamine (41 mg, 0.68 mmol). Themixture was stirred at 90° C. for 2 h. From this mixture the titlecompound was isolated using reverse phase prep HPLC. MS found forC₁₅H₂₀N₆O as (M+H)⁺ 301.2. UV λ=240, 292 nm.

Example 4174-(3-(2H-1,2,3-triazol-2-yOphenylamino)-2-(2-aminoethylamino)pyrimidine-5-carboxamide

Compound 179.6 (50 mg, 0.15 mmol) was dissolved in 3 mL NMP. To it wasadded MCPBA (49 mg, 0.18 mmol) and the mixture was stirred for 30 min.To it was added ethylenediamine (36 mg, 0.60 mmol). The mixture wasstirred at 90° C. for 2 h. From this mixture the title compound wasisolated using reverse phase prep HPLC. MS found for C₁₅H₁₇N₉O as (M+H)⁺340.3. UV λ=249 nm.

Example 4184-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(3-aminopropylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417. MS found for C₁₆H₁₉N₉O as (M+H)⁺ 354.4. UV λ=251 nm.

Example 4194-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(4-aminobutylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.4. UV λ=252 nm.

Example 4204-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(2-amino-2-methylpropylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.4. UV λ==249 nm.

Example 4214-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-((4-aminotetrahydro-2H-pyran-4-yl)methylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417. MS found for C₁₉H₂₃N₉O₂ as (M+H)⁺ 410.4. UV 2=249 nm.

Example 4224-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(2-(methylamino)ethylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 417 with commercially available tert-butyl2-aminoethyl(methyl)carbamate, followed by TFA treatment to cleave theBOC group. MS found for C₁₆H₁₉N₉O as (M+H)⁺ 354.4. UV λ=250 nm.

Example 4234-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1R,2S)-2-hydroxycyclohexylamino)pyrimidine-5-carboxamide(racemic)

The title racemic compound was prepared using the same chemistry shownfor Example 417 with commercially available cis-2-amino-1-cyclohexanoland DIEA. MS found for C₁₉H₂₂N₈O₂ as (M+H)⁺ 395.4. UV λ=254 nm.

Example 424(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-aminopropan-2-ylamino)pyrimidine-5-carboxamide

Step 1: Commercially available H-D-Ala-NH₂.HCl (2.0 g, 16 mmol) wasdissolved in 60 mL water and 20 mL dioxane. To it were added BzCl (2.7mL, 19.3 mmol) and sodium carbonate (5.1 g, 48 mmol). The mixture wasstirred for overnight at RT. It was diluted with ethyl acetate, washedwith brine x3, dried, concentrated. The residue was then taken into 450mL hexane and 50 mL DCM, stirred vigorously at 30° C. for 30 min,filtered. Compound 210.1 (>90% yield) stayed in the solid phase, andmost impurities and by-products were in the filtrate.

Step 2: Compound 210.1 (16 mmol) from Step 1 was dissolved in 100 mLTHF. To it was added BH₃.Me₂S (3.8 mL, 40 mmol) at RT. The mixture washeated to 85° C. and gently refluxed for 5 h. It was cooled to RT. To itwas added 100 mL water. The mixture was stirred for 1 h at RT. ThenK₂CO₃ (6.62 g, 48 mmol) and BOC₂O (7.00 g, 32 mmol) were added. Themixture was stirred at RT for 1 h. It was diluted with ethyl acetate,washed with brine x3, dried, concentrated, subjected to silica flashcolumn with 35% ethyl acetate in hexane to isolate compound 210.3 (1.22g, 25% for 3 steps overall) as white solid.

Step 3: Compound 210.3 (3.96 mmol) from Step 2 was dissolved in 200 mLethyl acetate. To it was added 500 mg 10% Pd/C and the mixture wasstirred under H₂ balloon overnight. It was filtered through celite, andthe celite was thoroughly rinsed with methanol. The filtrate wasconcentrated in vacuo to afford compound 210.4 as thick oil. It wasdissolved in NMP to make a 0.20M stocking solution.

Step 4: Compound 179.6 (100 mg, 0.30 mmol) was dissolved in 6 mL NMP. Toit was added MCPBA (96 mg, 0.36 mmol). The mixture was stirred for 40min at RT. TO it were added DIEA (0.21 mL, 1.2 mmol) and compound 210.4(0.20 M, 3 mL, 0.60 mmol). The mixture was stirred at 90° C. for 2 h. Itwas cooled to RT, diluted with ethyl acetate, washed with sat. Na₂CO₃x₂and brine, dried, concentrated in vacuo. The residue was treated withneat TFA at RT for 1 h and then concentrated. The residue was thensubjected to reverse phase prep HPLC to isolate the title compound. MSfound for C₁₆H₁₉N₉O as (M+H)⁺ 354.3. UV λ=250 nm.

Example 425(R)-2-(1-aminopropan-2-ylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the same chemistry scheme shown forExample 424 with compound 249.4 (given in Example 172). MS found forC₁₈H₂₀N₈O as (M+H)⁺ 365.3. UV λ=247 nm.

Example 426(R)-2-(1-aminopropan-2-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was made using the same chemistry scheme shown forExample 424 with compound 187.10 (given in Example 187). MS found forC₁₅H₂₀N₆₀ as (M+H)⁺ 301.3. UV λ=243, 289 nm.

Example 427(R)-2-(1-aminopropan-2-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was made using the same chemistry scheme shown forExample 424 with compound 185.1 (given in Example 195). MS found forC₁₆H₂₂N₆O as (M+H)⁺ 315.3. UV λ=243, 289 nm.

Example 428(S)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(2-aminopropylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₆H₁₉N₉O as (M+H)⁺ 354.3. UV λ=250 nm.

Example 215(S)-2-(2-aminopropylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₈H₂₀N₈O as (M+H)⁺ 365.3. UV λ=247 nm.

Example 216(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-3-methylbutan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₈H₂₃N₉O as (M+H)⁺ 382.4. UV λ=251 nm.

Example 217(R)-2-(1-amino-3-methylbutan-2-ylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₂₀H₂₄N₈O as (M+H)⁺ 393.4. UV λ=249 nm.

Example 218(S)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-3-hydroxypropan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₆H₁₉N₉O₂ as (M+H)⁺ 370.3. UV λ=248 nm.

Example 219(S)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-3-methoxypropan-2-ylamino)pyrimidine-5-carboxamide

Compound 219.1 was prepared using the same chemistry for compound 210.3shown in Example 414. Compound 219.1 (2.0 g, 6 mmol) was dissolved in100 mL DCM. To it were added proton sponge (3.2 g, 15 mmol) at RT. Threemin later, Me₃O⁺BF₄ ⁻ (2.2 g, 15 mmol) was added. The mixture wasstirred at RT for 3 days. It was diluted with 500 mL ethyl acetate,washed with brine x2, dried, concentrated, subjected to silica flashcolumn (40% ethyl acetate in hexane) to give compound 219.2 (1.77 g,87%) as white solid.

The title compound was completed using the similar chemistry schemeshown for Example 424 with compound 218.2. MS found for C₁₇H₂₁N₉O₂ as(M+H)⁺ 384.3. UV λ=249 nm.

Example 220(S)-2-(1-amino-3-methoxypropan-2-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 219. MS found for C₁₆H₂₂N₆O₂ as (M+H)⁺ 331.3. UV λ=240, 289 nm.

Example 221(R)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)-2-(1-amino-4-methylpentan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₉H₂₅N₉O as (M+H)⁺ 396.4. UV λ=250 nm.

Example 428(R)-4-(3-(1H-pyrazol-1-yl)phenylamino)-2-(1-amino-4-methylpentan-2-ylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₂₀H₂₆N₈O as (M+H)⁺ 395.4. UV λ=247 nm.

Example 429(R)-2-(1-amino-4-methylpentan-2-ylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₂₁H₂₆N₈O as (M+H)⁺ 405.4. UV 2=247 nm.

Example 431(R)-2-(1-amino-4-methylpentan-2-ylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₈H₂₆N₆O as (M+H)⁺ 343.4. UV λ=244, 288 nm.

Example 432(R)-2-(1-amino-4-methylpentan-2-ylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was made using the similar chemistry scheme shown forExample 424. MS found for C₁₉H₂₈N₆O as (M+H)⁺ 357.4. UV λ=241, 290 nm.

Example 4332-(4-aminobutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

To a solution of 1,4-diaminobutane (0.48 g, 5.5 mmol) in DMF (5 mL), asolution of compound2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-4-(m-tolylamino)pyrimidine-5-carboxamide(0.53 g, 1.47 mmol) in DMF (5 mL) was added dropwise. It was thenstirred at room temperature for 20 h. It was concentrated in vacuo. Theresidue was purified by HPLC to give the titled compound astrifluoroacetic acid salt (0.55 g). MS 315.2 (M+H); UV 249.7.

Example 434(S)-2-(2-amino-3-cyclopropylpropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₀H₂₅N₇O as (M+H)⁺ 380.4. UV:λ=219.8, 241.1, 330.3.

Example 435(S)-2-(2-aminobutylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₅N₇O as (M+H)⁺ 368.4. UV:λ=219.2, 240.4, 331.9.

Example 436(S)-2-(2-amino-2-cyclopropylethylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₃N₇O as (M+H)⁺ 366.4. UV:λ=220.2, 241.4, 330.6. NMR (CD₃OD): δ 8.27 (s, 1H), 8.08 (s, 1H), 7.85(d, J=2.4 Hz, 1H), 7.34 (s, 1H), 7.06-7.00 (m, 3H), 6.80 (m, 1H), 6.12(m, 1H), 4.18 (m, 1H), 2.67 (m, 2H), 1.19 (m, 3H), 0.97 (m, 1H),0.42-0.31 (m, 6H) ppm.

Example 437(S)-2-(2-aminobutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₃N₇O as (M+H)⁺ 354.4. UV:λ=219.2, 240.4, 333.1.

Example 438(R)-2-(2-amino-3-ethoxypropylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₀H₂₇N₇O as (M+H)⁺ 398.4. UV:λ=219.3, 239.4, 326.7.

Example 439(R)-2-(2-amino-3-methoxypropylamino)-4-(1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₁N₇O₂ as (M+H)⁺ 356.4.UV: λ=216.7, 238.7, 327.2.

Example 440(R)-2-(2-amino-3-methoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.4.UV: λ=219.2, 239.3, 331.9.

Example 441(R)-2-(2-amino-3-methylbutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₅N₇O as (M+H)⁺ 368.4. UV:λ=220.4, 325.9.

Example 4422-((2R,3R)-2-amino-3-methoxylbutylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₀H₂₇N₇O₂ as (M+H)⁺ 398.4.UV: λ=220.4, 239.9, 331.6.

Example 443(R)-2-(2-amino-3-ethoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.3.UV: λ=219.2, 239.3, 331.9.

Example 444(R)-2-(2-amino-3-ethoxypropylamino)-4-(1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.3.UV: λ=215.7, 238.1, 327.1.

Example 445(R)-2-(2-amino-3-methoxypropylamino)-4-(2-methyl-2H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₂N₈O₂ as (M+H)⁺ 371.3.UV: λ=214.9, 241.7, 324.1.

Example 446(R)-2-(2-amino-4-methoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.3.UV: λ=219.2, 240.4, 331.9.

Example 447(R)-2-(2-amino-3-methoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₅N₇O₂ as (M+H)⁺ 396.4.UV: λ=203.9, 244.0, 303.3.

Example 448(R)-2-(2-amino-3-ethoxypropylamino)-4-(2-methyl-2H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.4.UV: λ=208.6, 240.4, 283.1, 324.7.

Example 449(R)-2-(2-amino-3-ethoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4.UV: λ=221.6, 336.7.

Example 450(S)-2-(2-amino-4-methoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4.UV: λ=222.8, 336.7.

Example 4512-((2R,3R)-2-amino-3-methoxylbutylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4.UV: λ=221.6, 335.5.

Example 452(S)-2-(2-amino-3,3-dimethylbutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in

Example 218 MS found for C₂₀H₂₇N₇O as (M+H)⁺ 382.4. UV: λ=219.2, 239.3,327.1. Example 453(S)-2-(2-aminopropylamino)-4-(3-(3-fluorophenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide(41)

Step 1: Dichloropyrimidine ester 41.1 (3.44 g, 15.5 mmol) was dissolvedin acetonitrile (20 mL) and stirred in ice bath. To it was added asolution of aniline 41.2 (3.0 g, 14.1 mmol) and ethyldiisopropylamine(DIEA, 3.85 mL, 35.5 mmol) in 10 mL acetonitrile dropwise using anadditional funnel. The mixture was stirred for 1 hour, solventevaporated followed by dilution with water to precipitate compound 41.3as a yellow solid (4.2 g, 75%). MS: (M+H)⁺ 416.09

Step 2: Ethyl ester 41.3 (2 g, 5 mmol) was dissolved in 10 mL THF. To itwere added lithium hydroxide hydrate (275 mg, 6.5 mmol) and 10 mL water.The mixture was stirred for 1 hour and to it was carefully added 1N HClsolution till pH reaching 3. The mixture was concentrated in vacuo toremove THF. White solid crashed out and was isolated using a Büchnerfunnel. It was washed with water and dried in vacuum oven to givecompound 41.4 (1.65 g, 89%) as a white solid. MS: (M+H)⁺ 388.06

Step 3: Carboxylic acid 41.4 (1 g, 2.7 mmol) was dissolved in 15 mL DMF.To it were added EDC hydrochloride (934 mg, 4.86 mmol) and HOBt hydrate(660 mg, 4.86 mmol). The mixture was stirred at RT for 90 minutes. To itwas then added ammonia (commercial 0.5N solution in dioxane, 14 mL, 6.75mmol) at 0° C. The mixture was stirred for overnight at roomtemperature. To the reaction mixture added water to afford compound 41.5as a white solid (1.26 g, 86%). MS: (M+H)⁺ 485.14.

Step 4: Benzotriazolyl ether 41.5 (500 mg, 1 mmol) was dissolved in 15mL ACN. To it was added (5)-tert-butyl-1-aminopropane-2-yl-carbamate(373 mg, 2.1 mmol). The mixture was stirred for overnight at 50° C.After cooling added water and stirred for 1 hr to afford compound 41.6as a white solid (0.541 g, 92%). MS found for C₂₆H₃₀FN₇O₄ as (M+H)⁺524.24.

Step 5: Compound 41.6 (100 mg, 0.2 mmol) was stirred in a 1:1 (5 mL)mixture of TFA and dichloromethane at RT for 15 minutes. It wasconcentrated in vacuo followed by addition of diisopropyl ether toafford compound No. 41 as a white solid (50 mg, 59%). MS found forC₂₁H₂₂FN₇O₂ as (M+H)⁺ 424.19.

Example 454(S)-2-(2-aminopropylamino)-4-(3-(4-chlorophenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide(57)

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-(4-chlorophenyl)benzamide toreplace to 3-amino-N-(3-fluorophenyl)benzamide. MS found forC₂₁H₂₂ClN₇O₂ as (M+H)⁺ 440.

Example 455 (S)-2-(2-aminopropylamino)-4-(3-(isoxazol-3-ylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-(isoxazol-3-yl)benzamide toreplace to 3-amino-N-(3-fluorophenyl)benzamide. MS found for C₁₈H₂₀N₈O₃as (M+H)⁺ 396

Example 456 (S)-2-(2-aminopropylamino)-4-(3-(phenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-phenylbenzamide to replace to3-amino-N-(3-fluorophenyl)benzamide. MS found for C₂₁H₂₃N₇O₃ as (M+H)⁺406

Example 457(S)-2-(2-aminopropylamino)-4-(4-carbamoyl-3-methoxyphenylamino)pyrimidine-5-carboxamide(121)

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 4-amino-2-methoxybenzamide to replaceto 3-amino-N-(3-fluorophenyl)benzamide. MS found for C₁₆H₂₁N₇O₃ as(M+H)⁺ 360

Example 458(S)-2-(2-aminopropylamino)-4-(3-(5-methylthiazol-2-ylcarbamoyl)phenylamin)opyrimidine-5-carboxamide(123)

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with3-amino-N-(5-methylthiazol-2-yl)benzamide to replace to3-amino-N-(3-fluorophenyl)benzamide. MS found for C₁₉H₂₂N₈O₂S as (M+H)⁺427

Example 459(S)-2-(2-aminopropylamino)-4-(3-((5-chlorothiazol-2-ylmethylcarbamoyl)phenylamino)pyrimidine-5-carboxamide(125)

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with3-amino-N-((5-chlorothiazol-2-yl)methyl)benzamide to replace to3-amino-N-(3-fluorophenyl)benzamide. MS found for C₁₉H₂₁ClN₈O₂S as(M+H)⁺ 447

Example 460(S)-2-(2-aminopropylamino)-4-(3-(methoxyphenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-(4-methoxyphenyl)benzamide toreplace to 3-amino-N-(3-fluorophenyl)benzamide. MS found for C₂₂H₂₅N₇O₃as (M+H)⁺ 436

Example 461(S)-2-(2-aminopropylamino)-4-(3-(3-chlorophenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-(3-chlorophenyl)benzamide toreplace to 3-amino-N-(3-fluorophenyl)benzamide. MS found forC₂₁H₂₂ClN₇O₂ as (M+H)⁺ 440

Example 462S)-2-(2-aminopropylamino)-4-(3-(4-fluorophenylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with 3-amino-N-(4-fluoroophenyl)benzamide toreplace to 3-amino-N-(3-fluorophenyl)benzamide. MS found for C₂₁H₂₂FN₇O₂as (M+H)⁺ 424

Example 463S)-2-(2-aminopropylamino)-4-(3-(benzo[d]thiazol-2-ylcarbamoyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 453 with3-amino-N-(benzo[d]thiazol-2-yl)benzamide to replace to3-amino-N-(3-fluorophenyl)benzamide. MS found for C₂₂H₂₂N₈O₂S as (M+H)⁺463

The compounds No. 140, 141, 144, 156, 163, 164, 166, 172, 173, 176, 177,180, 191 194, 200, 201, 213, 215, 218, 221, 222, 228, 233, 237, 238, and239 were prepared using the same synthetic scheme demonstrated inExample 453.

Example 464(S)-2-(2-aminopropylamino)-4-(4-methyl-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was made in an analogous fashion to 81 in Example 87.However, instead of the 4-(thiazol-4-yl) derivative 81.1, the4-methyl-3-(2H-1,2,3-triazol-2-yl) derivative was utilized. Also,instead of tert-butyl (1S,2R)-2-aminocyclohexylcarbamate 81.2,N—((S)-2-amino-1-methylethyl)carbamic acid tert-butyl ester 333.2 [CAS146552-71-8] was used. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.4. UV λ=242nm.

Example 465(S)-2-(2-aminopropylamino)-4-(5-methyl-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

This compound was made in an analogous fashion to 333 in example 464.However, instead of the 4-methyl-3-(2H-1,2,3-triazol-2-yl) derivative333.1, the 5-methyl-3-(2H-1,2,3-triazol-2-yl) derivative 334.1 wasutilized. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.3. UV λ=246 nm

Example 466(S)-2-(2-aminopropylamino)-4-(3-(2H-1,2,3-triazol-2-yl)-5-(trifluoromethyl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared according the scheme 464 using 335.3 asthe starting aniline. MS found for C₁₇H₁₈F₃N₉O as (M+H)⁺ 422.3. UV λ=260nm

Example 467(R)-2-(1-aminobutan-2-ylamino)-4-(3-methylphenyl)pyrimidine-5-carboxamide

Commercially available (R)-(−)-2-butanamide hydrochloride (346.1; 2.41g, 17.4 mmol) [CAS 103765-03-3] was dissolved in water/dioxane (60 mL/20mL) containing Na₂CO₃ (5.53 g, 52.2 mmol). To this stirring solution wasadded benzyl chloroformate (2.81 mL, 20.9 mmol) [CAS 501-53-1]. Thereaction was stirred for 96 h at RT. The reaction mixture was dilutedwith EtOAc (400 mL) and water (100 mL). The layers were separated andthe organics were further washed with water (2×150 mL) and brine (2×150mL). The EtOAc layer was dried (MgSO₄) and concentrated. The resultingwhite solid was triturated with 360 mL hexanes/50 mL CH₂Cl₂ for 1 h. Thesolid was filtered, washed with hexanes, and dried to give 346.2 (3.46g).

To a stirring solution of 346.2 (3.46 g, 14.7 mmol) in anhydrous THF(100 mL) was added BH₃.Me₂S (3.45 mL, 36.6 mmol) dropwise. The resultingsolution was heated under an argon atmosphere at 85° C. for 18 h. Thereaction mixture was cooled and ice-cold water was slowly added (100mL). The resulting solution was stirred for 1 h. Next, K₂CO₃ was added(6.08 g, 44.1 mmol) followed by Boc anhydride (6.40 g, 29.4 mmol). Thereaction was stirred for 18 h and then diluted with EtOAc. The organiclayer was washed with 5% K₂CO₃, 5% NaHCO₃, water, and brine. It was thendried over MgSO₄ and concentrated to yield 346.3.

Crude 346.3 was dissolved in ˜50 mL EtOAc and a catalytic amount of 10%Pd/C was added. To this suspension was amounted a hydrogen balloon forovernight stirring. The mixture was filtered through celite andconcentrated in vacuo to afford 346.4.

Intermediate 346.4 was reacted with 344.1 according to the chemistrydescribed in example 13. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₆H₂₂N₆O as(M+H)⁺ 315.4. UV λ=238, 284 nm.

Example 468(R)-2-(1-aminobutan-2-ylamino)-4-(3,5-dimethylphenyl)pyrimidine-5-carboxamide

Intermediate 346.4 was reacted with 345.1 according to the chemistrydescribed in example 13. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₇H₂₄N₆O as(M+H)⁺ 329.5. UV λ=240, 290 nm. δ 1.05 (t, 3H), 1.70-1.80 (m, 2H), 2.38(s, 6H), 3.15-3.35 (m, 2H), 4.16-4.23 (m, 1H), 6.90 (s, 1H), 7.19-7.25(m, 2H), 8.50 (s, 1H)

Example 469 (R)-2-(1-aminobutan-2-ylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Intermediate 346.4 was reacted with 343.5 according to the chemistrydescribed in Example 424. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₇H₂₁N₉O as(M+H)⁺ 368.4. UV λ=248 nm. δ 1.05 (t, 3H), 1.70-1.80 (m, 2H), 3.10-3.35(m, 2H), 4.55-4.61 (m, 1H), 7.35 (d, 1H), 7.55 (t, 1H), 7.92 (d, 1H),8.00 (s, 2H), 8.58 (s, 1H), 9.00 (s, 1H)

Example 470(R)-2-(1-aminobutan-2-ylamino)-4-(3-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

Intermediate 346.4 was reacted with 349.1 according to the chemistrydescribed in Example 424. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₉H₂₂N₈O as(M+H)⁺ 379.5. UV λ=248 nm. δ 1.05 (t, 3H), 1.70-1.80 (m, 2H), 3.10-3.35(m, 2H), 4.48-4.55 (m, 1H), 7.41 (t, 1H), 7.55-7.60 (m, 2H), 8.25-8.35(m, 1H), 8.55 (s, 1H), 8.89 (d, 2H), 9.00 (s, 1H)

Example 4712-((R)-1-aminobutan-2-ylamino)-4-(3-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

Intermediate 346.4 was reacted with 350.1 according to the chemistrydescribed in Example 424. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₈H₂₂N₈O as(M+H)⁺ 367.4. UV λ=246 nm. δ 1.05 (t, 3H), 1.70-1.80 (m, 2H), 3.10-3.35(m, 2H), 4.48-4.55 (m, 1H), 6.58 (br s, 1H), 7.30 (d, 1H), 7.50-7.62 (m,2H), 7.80 (br s, 1H), 8.32 (br s, 1H), 8.55 (s, 1H), 8.68 (br s, 1H)

Example 472(R)-2-(1-amino-3-cyclopropylpropan-2-ylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Intermediate 354.5 was reacted with 343.5 according to the chemistrydescribed in Example 424. After Cbz-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₉H₂₃N₉O as(M+H)⁺ 394.4. UV λ=250 nm.

Example 473(R)-2-(1-amino-3-cyclopropylethylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was made in an analogous fashion to Example 424. MSfound for C₁₈H₂₁N₉O as (M+H)⁺ 380.4. UV λ=250 nm. δ 0.50-0.85 (m, 4H),1.15-1.25 (m, 1H), 3.10-3.35 (m, 2H), 3.90-4.00 (m, 1H), 7.30-7.36 (m,1H), 7.60 (t, 1H), 7.93-8.00 (m, 1H), 8.05 (s, 2H), 8.56 (s, 1H), 8.90(br s, 1H).

Example 4742-((1-aminocyclopropylmethyl)amino)-4-(3-(2H-1,2,3-triazol-2-yOphenylamino)pyrimidine-5-carboxamide

The title compound was made in an analogous fashion to Example 424. MSfound for C₁₇H₁₉N₉O as (M+H)⁺ 366.4. UV λ=248 nm.

Example 4752-((1-aminocyclopropylmethyl)amino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was made in an analogous fashion to Example 424. MSfound for C₁₈H₂₁N₇O as (M+H)⁺ 352.4. UV 2=217, 239, 331 nm.

Example 4762-((1-aminocyclopropylmethyl)amino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was made in an analogous fashion to Example 424. MSfound for C₁₇H₂₂N₆O as (M+H)⁺ 327.4. UV λ=241, 290 nm.

Example 4782-(1-amino-4,4,4-trifluorobutan-2-ylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Intermediate 360.6 was reacted with 343.5 according to the chemistrydescribed in Example 424. After Boc-deprotection, reverse phasepreparative HPLC afforded the title compound. MS found for C₁₇H₁₈F₃N₉Oas (M+H)⁺ 422.4. UV λ=245 nm. 2.70-2.80 (m, 2H), 4.25-4.35 (m, 1H),7.35-7.41 (m, 1H), 7.49-7.55 (m, 1H), 7.88-7.94 (m, 1H), 7.98 (s, 2H),8.59 (s, 1H), 8.86 (br s, 1H).

Example 4792-((1-methylaminocyclopropylmethyl)amino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

Intermediate 345.1 was reacted with1-(aminomethyl)-N-methylcyclopropanamine according to Example 408.Reverse phase preparative HPLC afforded the title compound. MS found forC₁₈H₂₄N₆O as (M+H)⁺ 341.5. UV 2=240 nm.

Example 480(S)-2-(2-amino-3-cyclopropylpropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₀H₂₅N₇O as (M+H)⁺ 380.4. UV:λ=219.8, 241.1, 330.3.

Example 481(S)-2-(2-aminobutylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₅N₇O as (M+H)⁺ 368.4. UV:λ=219.2, 240.4, 331.9.

Example 482(S)-2-(2-amino-2-cyclopropylethylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₃N₇O as (M+H)⁺ 366.4. UV:λ=220.2, 241.4, 330.6.

Example 483(S)-2-(2-aminobutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₃N₇O as (M+H)⁺ 354.4. UV:λ=219.2, 240.4, 333.1.

Example 484(R)-2-(2-amino-3-ethoxypropylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₇N₇O as (M+H)⁺ 398.4. UV:λ=219.3, 239.4, 326.7.

Example 485(R)-2-(2-amino-3-methoxypropylamino)-4-(1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₁N₇O₂ as (M+H)⁺ 356.4.UV: λ=216.7, 238.7, 327.2.

Example 486(R)-2-(2-amino-3-methoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxnmide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.4.UV: λ=219.2, 239.3, 331.9.

Example 487(R)-2-(2-amino-3-methylbutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₅N₇O as (M+H)⁺ 368.4. UV:λ=220.4, 325.9.

Example 4882-((2R,3R)-2-amino-3-methoxylbutylamino)-4-(1-ethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₇N₇O₂ as (M+H)⁺ 398.4.UV: λ=220.4, 239.9, 331.6.

Example 489(R)-2-(2-amino-3-ethoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.3.UV: λ=219.2, 239.3, 331.9.

Example 490(R)-2-(2-amino-3-ethoxypropylamino)-4-(1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.3.UV: λ=215.7, 238.1, 327.1.

Example 491(R)-2-(2-amino-3-methoxypropylamino)-4-(2-methyl-2H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₂N₈O₂ as (M+H)⁺ 371.3.UV: λ=214.9, 241.7, 324.1.

Example 492(R)-2-(2-amino-4-methoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.3.UV: λ=219.2, 240.4, 331.9.

Example 493(R)-2-(2-amino-3-methoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₅N₇O₂ as (M+H)⁺ 396.4.UV: λ=203.9, 244.0, 303.3.

Example 494(R)-2-(2-amino-3-ethoxypropylamino)-4-(2-methyl-2H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.4.UV: λ=208.6, 240.4, 283.1, 324.7.

Example 495(R)-2-(2-amino-3-ethoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4.UV: λ=221.6, 336.7.

Example 496(S)-2-(2-amino-4-methoxypropylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in

Example 596 MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4. UV: λ=222.8, 336.7.Example 4972-((2R,3R)-2-amino-3-methoxylbutylamino)-4-(2,3-dihydro-1H-pyrrolo[1,2-a]indol-8-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₇N₇O₂ as (M+H)⁺ 410.4.UV: λ=221.6, 335.5.

Example 498(S)-2-(2-amino-3,3-dimethylbutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₀H₂₇N₇O as (M+H)⁺ 382.4. UV:λ=219.2, 239.3, 327.1.

Example 499(R)-2-(2-amino-3-methoxypropylamino)-4-(6-carbomoylnaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₃N₇O₃ as (M+H)⁺ 410.2.UV: λ=224.9, 316.2.

Example 500(R)-2-(2-amino-3-methoxypropylamino)-4-(1,2-dimethyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.4.UV: λ=223.9.

Example 5012-((2R,3R)-2-amino-3-methoxybutylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.3.UV: λ=203.9, 241.6.

Example 502(R)-2-(2-amino-3-methoxypropylamino)-4-(2-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.4.UV: λ=220.4, 333.1.

Example 503(S)-2-(2-amino-4-methoxybutylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.4.UV: λ=209.8, 240.4, 283.1, 325.9.

Example 504(S)-2-(2-amino-3-cyclopropylpropylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₄N₆O as (M+H)⁺ 341.4. UV:λ=241.7, 285.9.

Example 5052-((2R,3R)-2-amino-3-methoxybutylamino)-4-(1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₃N₇O₂ as (M+H)⁺ 370.4.UV: λ=216.7, 238.1, 324.8.

Example 506(R)-2-(2-amino-3-methoxypropylamino)-4-(naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₂N₆O₂ as (M+H)⁺ 367.2.UV: λ=213.3, 239.3, 305.6.

Example 507(R)-2-(2-amino-3-ethoxypropylamino)-4-(2-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.4.UV: λ=220.4, 240.4.

Example 508(R)-2-(2-amino-3-phenoxypropylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₃H₂₅N₇O₂ as (M+H)⁺ 432.4.UV: λ=218.6, 242.3.

Example 509(S)-2-(2-amino-4-methylpentylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₇N₇O as (M+H)⁺ 382.4. UV:λ=219.8, 241.7.

Example 510(R)-2-(2-amino-3-methoxypropylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.4.UV: λ=240.3, 290.0.

Example 5112-((2R,3R)-2-amino-3-methoxybutylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₅N₇O₂ as (M+H)⁺ 384.4.UV: λ=218.0

Example 512(R)-2-(2-amino-3-phenoxypropylamino)-4-(2-methyl-2H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₂H₂₅N₈O₂ as (M+H)⁺ 433.4.UV: λ=211.3, 242.3, 330.9.

Example 513(R)-2-(2-amino-3-methoxypropylamino)-4-(biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₄N₆O₂ as (M+H)⁺ 393.4.UV: λ=202.8, 246.3.

Example 514(R)-2-(2-amino-3-methoxypropylamino)-4-(6-methoxynaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₄N₆O₃ as (M+H)⁺ 397.0.UV: λ=221.6, 315.2.

Example 515(R)-2-(2-amino-3-phenoxypropylamino)-4-(biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₆H₂₆N₆O₂ as (M+H)⁺ 455.4.UV: λ=203.4, 246.6.

Example 516(R)-2-(2-amino-3-methoxypropylamino)-4-(6-(methylcarbamoylnaphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in

Example 596 MS found for C₂₁H₂₅N₇O₃ as (M+H)⁺ 424.4. UV: λ=222.8, 316.4.Example 517(R)-2-(2-amino-3-methoxypropylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₋₁₇H₂₂N₈O₂ as (M+H)⁺ 371.5.UV: λ=208.8, 241.7, 316.7.

Example 518(S)-2-(2-aminobutylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₄N₆O as (M+H)⁺ 329.5. UV:λ=240.5, 288.4.

Example 519(S)-2-(2-aminobutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₆H₂₂N₆O as (M+H)⁺ 315.4. UV:λ=241.1, 286.5.

Example 520(R)-2-(2-amino-3-methoxypropylamino)-4-(1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₂₀N₈O₂ as (M+H)⁺ 357.3.UV: λ=206.3, 240.4, 311.6.

Example 521(R)-2-(2-amino-3-methoxypropylamino)-4-(3,5-dimethoxyphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₄ as (M+H)⁺ 377.3.UV: λ=203.8, 235.5, 290.0.

Example 522(R)-2-(2-amino-3-methoxypropylamino)-4-(6-(morpholine-4-carbonyl)naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₄H₂₉N₇O₄ as (M+H)⁺ 480.4.UV: λ=218.0, 311.6.

Example 5232-amino-3-(5-carbamoyl)-4-(m-tolylamino)pyrimidin-2-ylamino)propanoicacid

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₅H₁₈N₆O₃ as (M+H)⁺ 331.1.UV: λ=239.3, 289.0.

Example 524(S)-2-(2-amino-3-cyclopropylpropylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₆N₆O as (M+H)⁺ 355.5. UV:λ=241.1, 287.7.

Example 525(S)-2-(2-amino-3-methylbutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₄N₆O as (M+H)⁺ 329.4. UV:λ=241.7, 285.9.

Example 526(R)-2-(2-amino-3-methoxypropylamino)-4-(naphthalen-1-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₋₁₉H₂₂N₆O₂ as (M+H)⁺ 367.4.UV: λ=219.2, 282.8.

Example 527(S)-2-(2-amino-2-cyclopropylethylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₂N₆O as (M+H)⁺ 327.4. UV:λ=241.7, 286.5.

Example 528(R)-2-(2-amino-3-methoxypropylamino)-4-(6-(dimethylcarbamoyl)naphthalen-2-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₂H₂₇N₇O₃ as (M+H)⁺ 438.2.UV: λ=216.9, 309.2.

Example 5292-((2R,3R)-2-amino-3-methoxybutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.4.UV: λ=240.4, 284.2.

Example 530(S)-2-(2-amino-2-cyclopropylethylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₄N₆O as (M+H)⁺ 341.4. UV:λ=241.4, 287.7.

Example 531(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(pyridine-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₃N₇O₂ as (M+H)⁺ 394.4.UV: λ=242.8.

Example 532(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(thiazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₁N₇O₂S as (M+H)⁺ 400.2.UV: λ=240.4, 290.2.

Example 533(S)-2-(2-amino-3-cyclopropylpropylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₃N₉O as (M+H)⁺ 394.4. UV:λ=204.6, 251.5.

Example 534(R)-2-(2-amino-3-methoxypropylamino)-4-(3,4-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.5.UV: λ=239.3, 291.4.

Example 535(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(isoxazol-3-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₁N₇O₃ as (M+H)⁺ 384.3.UV: λ=242.3, 285.4.

Example 536(S)-3-amino-4-(5-carbamoyl)-4-(m-tolylamino)pyrimidin-2-ylamino)butanoicacid

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₆H₂₀N₆O₃ as (M+H)⁺ 345.1.UV: λ=239.3, 285.4.

Example 537(R)-2-(2-amino-3-ethoxypropylamino)-4-(1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₂N₈O₂ as (M+H)⁺ 371.3.UV: λ=202.8, 238.1, 308.0.

Example 538(R)-2-(2-amino-3-ethoxypropylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.3.UV: λ=208.6, 240.4, 319.9.

Example 539(R)-2-(2-aminobutylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₁N₉O as (M+H)⁺ 368.4. UV:λ=203.9, 249.9.

Example 540(S)-2-(2-amino-4-methoxybutylamino)-4-(biphenyl-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₂H₂₆N₆O₂ as (M+H)⁺ 407.4.UV: λ=202.1, 246.0.

Example 541(S)-2-(2-amino-3,3-dimethylbutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₆N₆O as (M+H)⁺ 343.4. UV:λ=240.5.

Example 542 (R)-2-(2-amino-3-methoxypropylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₁N₇O₂S as (M+H)⁺ 374.4.UV: λ=238.7, 295.1.

Example 543(R)-2-(2-amino-3-methoxypropylamino)-4-(4-chloro-3-methylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₂₁IN₆O₂ as (M+H)⁺ 365.2,367.1 (Cl pattern). UV: λ=215.7, 240.4, 289.0.

Example 544(R)-2-(2-amino-3-ethoxypropylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.6.UV: λ=238.1, 284.2.

Example 545(R)-2-(2-amino-3-isopropoxypropylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₆N₆O₂ as (M+H)⁺ 359.8.UV: λ=241.6, 286.6.

Example 546(S)-2-(2-amino-3-methylbutylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₄N₆O as (M+H)⁺ 343.5. UV:λ=239.9, 284.7.

Example 5472-((2R,3R)-2-amino-3-methoxybutylamino)-4-(1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₂N₈O₂ as (M+H)⁺ 371.4.UV: λ=238.1, 299.7.

Example 548(R)-2-(2-amino-3-methoxypropylamino)-4-(3-methoxyphenyl)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₂₂N₆O₃ as (M+H)⁺ 347.2.UV: λ=239.7.

Example 549(R)-2-(2-amino-3-methoxypropylamino)-4-(3-ethylphenyl)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.2.UV: λ=239.3, 286.6.

Example 550(S)-2-(2-amino-4-(fluorophenyl)propylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₃H₂₄FN₇O as (M+H)⁺ 434.5.UV: λ=215.5, 241.7, 330.9.

Example 551(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₁N₉O₂ as (M+H)⁺ 384.3.UV: λ=249.9.

Example 552(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(1H-pyrazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.3.UV: λ=247.2.

Example 553(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(pyrimidin-5-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₋₁₉H₂₂N₈O₂ as (M+H)⁺ 395.3.UV: λ=243.6.

Example 554(R)-2-(2-amino-3-hydroxypropylamino)-4-(m-tolyl)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₅H₂₀N₆O₂ as (M+H)⁺ 317.2.UV: λ=238.1, 286.6.

Example 555(R)-4-(1-methyl-1H-indol-4-ylamino)-2-(piperidin-3-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₃N₇O as (M+H)⁺ 366.4. UV:λ=218.0, 241.6, 335.5.

Example 556(R)-2-(2-amino-3-hydroxy-3-methylbutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.2.UV: λ=239.3.

Example 557(S)-2-(2-amino-4-methoxybutylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.4.UV: λ=240.5, 286.5.

Example 5584-(1-methyl-1H-indol-4-ylamino)-2-(2-(methylamino)ethylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₇H₂₁N₇O as (M+H)⁺ 340.4. UV:λ=220.4, 239.3.

Example 5592-((2R,3S)-2-amino-3-methoxybutylamino)-4-(m-tolyl)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 345.2.UV: λ=239.3, 285.4.

Example 5602-((2R,3S)-2-amino-3-methoxybutylamino)-4-(3-methoxyphenyl)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₃ as (M+H)⁺ 361.2.UV: λ=239.3.

Example 561(R)-2-(2-amino-3-methoxypropylamino)-4-(3-(1H-imidazol-1-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₂N₈O₂ as (M+H)⁺ 383.5.UV: λ=238.1, 281.9.

Example 562(R)-2-(1-amino-4-methyl-1-thioxopentan-2-ylamino)-4-(1-methyl-1H-indol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 318. MS found for C₂₀H₂₅N₇OS as (M+H)⁺ 412.4.UV: λ=250.3.

Example 5632-((2R,3R)-2-amino-3-methoxybutylamino)-4-(1-methyl-1H-indazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in

Example 596 MS found for C₁₈H₂₄N₈O₂ as (M+H)⁺ 385.3. UV: λ=208.6, 240.4,311.6. Example 564(R)-2-(2-amino-3-methoxypropylamino)-4-(4-methoxy-3-methylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₋₁₇H₂₄N₆O₃ as (M+H)⁺ 361.2.UV: λ=238.1, 292.6.

Example 565(R)-2-(2-amino-3-methoxypropylamino)-4-(benzo[d]thiazol-7-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₁₉N₇O₂S as (M+H)⁺ 374.3.UV: λ=232.6, 287.8.

Example 566(S)-2-(2-amino-4-methylpentylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₈N₆O as (M+H)⁺ 357.4. UV:λ=239.3, 287.8.

Example 567(S)-2-(2-amino-2-cyclopropylethylamino)-4-(3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₁N₉O as (M+H)⁺ 380.4. UV:λ=204.6, 250.9.

Example 568(R)-2-(2-amino-3-methoxypropylamino)-4-(4-fluoro-3-methylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₂₁FN₆O₂ as (M+H)⁺ 349.0.UV: λ=211.0, 239.3.

Example 569(R)-2-(2-amino-3-methoxypropylamino)-4-(3-trifluoromethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₁₉F₃N₆O₂ as (M+H)⁺ 385.2.UV: λ=242.8.

Example 570 Methyl2-amino-3-(5-carbamoyl)-4-(m-tolylamino)pyrimidin-2-ylamino)propanoate

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₆H₂₀N₆O₃ as (M+H)⁺ 345.1.UV: λ=238.1, 290.2.

Example 580(R)-2-(2-amino-3-phenoxypropylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₁H₂₄N₆O₂ as (M+H)⁺ . . . UV:λ=213.3, 239.3.

Example 581 (R)-2-(2-amino-3-methoxypropylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₆H₁₉N₇O₂S as (M+H)⁺ 374.3.UV: λ=243.6, 292.1.

Example 582(S)-2-(2-amino-4-methylpentylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₆N₆O as (M+H)⁺ 343.4. UV:λ=239.3, 285.4.

Example 583(R)-2-(2-amino-3-methoxypropylamino)-4-(3,4-dimethoxyphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₄ as (M+H)⁺ 377.1.UV: λ=235.7, 285.4.

Example 584(S)-2-(2-amino-3-(thiazol-4-yl)propylamino)-4-(m-tolylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₈H₂₁N₇OS as (M+H)⁺ 384.1.UV: λ=240.4, 287.8.

Example 585(R)-2-(2-amino-3-methoxypropylamino)-4-(benzo[c][1,2,5]thiadiazol-4-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₅H₁₈N₈O₂S as (M+H)⁺ 375.2.UV: λ=235.7, 315.2.

Example 586(R)-2-(2-amino-3-methoxypropylamino)-4-(4-(pyrimidin-2-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₉H₂₂N₈O₂ as (M+H)⁺ 395.4.UV: λ=230.1, 311.2.

Example 587(R)-2-(2-amino-3-methoxypropylamino)-4-(quinoxalin-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₀N₈O₂ as (M+H)⁺ 369.2.UV: λ=245.4.

Example 588(R)-2-(2-amino-3-methoxypropylamino)-4-(4-(dimethylamino)-3-methylphenylylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₇N₇O₂ as (M+H)⁺ 374.3.

Example 589(R)-2-(2-amino-3-methoxypropylamino)-4-(4-(2-methoxyethoxy)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₈H₂₆N₆O₄ as (M+H)⁺ 391.4.UV: λ=202.8, 239.9, 282.2.

Example 590(R)-2-(2-amino-3-methoxypropylamino)-4-(3-methyl-4-morpholinophenylylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₂₀H₂₉N₇O₃ as (M+H)⁺ 416.2.UV: λ=238.1, 293.8.

Example 591(R)-2-(2-amino-3-methoxypropylamino)-4-(2,3-dihydrobenzo[b][1,4]dioxin-6-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₂N₆O₄ as (M+H)⁺ 375.3.UV: λ=239.3, 292.7.

Example 592(S)-2-(2-amino-3,3-dimethylbutylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₈N₆O as (M+H)⁺ 357.4. UV:λ=240.4.

Example 593(S)-2-(2-amino-4-(fluorophenyl)propylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₂₂H₂₅FN₆O as (M+H)⁺ 409.4.UV: λ=241.7, 288.4.

Example 594(R)-2-(2-amino-3-methoxypropylamino)-4-(benzo[d]isoxazol-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₋₁₆H₁₉N₇O₃ as (M+H)⁺ 358.3.UV: λ=203.9, 235.7, 292.6.

Example 595(R)-2-(2-amino-3-hydroxy-3-methylbutylamino)-4-(3,5-dimethylphenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 596. MS found for C₁₇H₂₄N₆O₂ as (M+H)⁺ 359.2.UV: λ=239.3.

Example 596

Synthesis of 243.2: To a suspension of NaH (6 g, 0.25 mol) in THF (220mL) at 0° C. was added MeOH (18 mL) dropwise, the mixture was thenstirred at ambient temperature for 1 h, and the resulting solution wasused next.

To a solution of N-Boc L-serine 243.1 (6 g, 0.029 mol) in THF (300 mL)was added 120 mL of above solution and MeI (3 mL). After stirring atambient temperature for 1 h, the remaining above solution was added,followed by more MeI (6 mL), and the mixture was stirred for additional18 h. The mixture was then concentrated under vacuum to remove THF, theresidue was dissolved in water; the aqueous solution was washed withether, and acidified with citric acid to pH 2. The acidified aqueoussolution was extracted with EtOAc (3×), EtOAc layer was combined, washedwith diluted Na₂S₂O₃, brine, dried and concentrated to give a mixture of243.2 and recovered 243.1. The residue was taken up in H₂O (100 mL), andwas extracted with DCM (3×). DCM layer was combined, washed with brine,dried over Na₂SO₄, and was concentrated in vacuo to give 243.2 as paleyellow oil (3.2 g).

Synthesis of 243.3: To a solution of 243.2 (3.2 g, 14.60 mmol) in THF(20 mL) at −15° C. was added N-methyl Morpholine (1.60 mL, 14.60 mmol)and isobutyl chloroformate (1.91 mL, 14.60 mmol). After stirring for 5min at −15° C., the mixture was added a solution of Sodium Borohydride(1.66 g, 43.8 mmol) in water (7 mL) slowly (gas evolution). The mixturewas kept stirring at −15° C. for 10 min, and was diluted with EtOAc, theorganics were washed with water, Sat. NaHCO₃, brine, dried andconcentrated to give crude residue, which was purified by column(Hex/EtOAc=2:1 to 2:3) to give 243.3 (2.1 g, 70%).

Synthesis of 243.4: To a solution of 243.3 (2.1 g, 10.24 mmol) in DCM(30 mL) at 0° C. was added Triethylamine (2.15 mL, 15.36 mmol) andMethanesulfonyl Chloride (1.19 mL, 15.36 mmol). After stirring at 0° C.for 30 min, it was diluted with DCM, the organics were washed withwater, Sat. NaHCO₃, brine, dried and concentrated to give crudemesylate.

To a solution of the above crude mesylate in DMF (20 mL) was added NaN₃(1.95 g, 30.72 mmol) and TBAI (37 mg, 0.124 mmol). After heating at 75°C. for 2 h, it was diluted with EtOAc, the organics were washed withwater, brine, dried and concentrated to give crude azide (1.7 g).

To a solution of the above crude azide (1.7 g) in EtOAc (16 mL) wasadded Pd/C (400 mg), and was charged with H₂ (1 atm). After stirring atambient temperature for 15 h, Pd/C was filtered off, and the filtratewas concentrated to give 243.4 (1.5 g).

Synthesis of 243: To a solid sample of 243.5 (0.76 g, 2.1 mmol) insealed tube was added a solution of 243.4 (0.75 g, 3.68 mmol) in AcCN(8.5 mL). After heating at 65° C. for 5 h, the mixture was cooled andadded water (40 mL). The precipitates were collected by filtration togive coupling product (0.85 g).

To a suspension of the above coupling product (0.85 g) in DCM (6 mL) wasadded TFA (2 mL). After stirring at ambient temperature for 15 min, themixture was concentrated, and the residue was purified by preparativeHPLC to give F (0.99 g) as TFA salt.

Example 597

Synthesis of 243.2: To a suspension of NaH (6 g, 0.25 mol) in THF (220mL) at 0° C. was added MeOH (18 mL) dropwise, the mixture was thenstirred at ambient temperature for 1 h, and the resulting solution wasused next.

To a solution of N-Boc L-serine 243.1 (6 g, 0.029 mol) in THF (300 mL)was added 120 mL of above solution and MeI (3 mL). After stirring atambient temperature for 1 h, the remaining above solution was added,followed by more MeI (6 mL), and the mixture was stirred for additional18 h. The mixture was then concentrated under vacuum to remove THF, theresidue was dissolved in water; the aqueous solution was washed withether, and acidified with citric acid to pH 2. The acidified aqueoussolution was extracted with EtOAc (3×), EtOAc layer was combined, washedwith diluted Na₂S₂O₃, brine, dried and concentrated to give a mixture of243.2 and recovered 243.1. The residue was taken up in H₂O (100 mL), andwas extracted with DCM (3×). DCM layer was combined, washed with brine,dried over Na₂SO₄, and was concentrated in vacuo to give 243.2 as paleyellow oil (3.2 g).

Synthesis of 243.3: To a solution of 243.2 (3.2 g, 14.60 mmol) in THF(20 mL) at −15° C. was added N-methyl Morpholine (1.60 mL, 14.60 mmol)and isobutyl chloroformate (1.91 mL, 14.60 mmol). After stirring for 5min at −15° C., the mixture was added a solution of Sodium Borohydride(1.66 g, 43.8 mmol) in water (7 mL) slowly (gas evolution). The mixturewas kept stirring at −15° C. for 10 min, and was diluted with EtOAc, theorganics were washed with water, Sat. NaHCO₃, brine, dried andconcentrated to give crude residue, which was purified by column(Hex/EtOAc=2:1 to 2:3) to give 243.3 (2.1 g, 70%).

Synthesis of 243.4: To a solution of 243.3 (2.1 g, 10.24 mmol) in DCM(30 mL) at 0° C. was added Triethylamine (2.15 mL, 15.36 mmol) andMethanesulfonyl Chloride (1.19 mL, 15.36 mmol). After stirring at 0° C.for 30 min, it was diluted with DCM, the organics were washed withwater, Sat. NaHCO₃, brine, dried and concentrated to give crudemesylate.

To a solution of the above crude mesylate in DMF (20 mL) was added NaN₃(1.95 g, 30.72 mmol) and TBAI (37 mg, 0.124 mmol). After heating at 75°C. for 2 h, it was diluted with EtOAc, the organics were washed withwater, brine, dried and concentrated to give crude azide (1.7 g).

To a solution of the above crude azide (1.7 g) in EtOAc (16 mL) wasadded Pd/C (400 mg), and was charged with H₂ (1 atm). After stirring atambient temperature for 15 h, Pd/C was filtered off, and the filtratewas concentrated to give 243.4 (1.5 g).

Synthesis of 243: To a solid sample of 243.5 (0.76 g, 2.1 mmol) insealed tube was added a solution of 243.4 (0.75 g, 3.68 mmol) in AcCN(8.5 mL). After heating at 65° C. for 5 h, the mixture was cooled andadded water (40 mL). The precipitates were collected by filtration togive coupling product (0.85 g).

To a suspension of the above coupling product (0.85 g) in DCM (6 mL) wasadded TFA (2 mL). After stirring at ambient temperature for 15 min, themixture was concentrated, and the residue was purified by preparativeHPLC to give F (0.99 g) as TFA salt.

Example 598(S)-2-(2-amino-2-cyclopropylethylamino)-4-(2-amino-3,3-dimethylbutylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₉H₂₅N₉O as (M+H)⁺ 396.4. UV:λ=248.7.

Example 5992-(1-acetylpiperidin-4-ylamino)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example 218. MS found for C₁₅H₂₂N₆O₂ as (M+H)⁺ 319.4.UV: λ=238.7.

Example 600(S)-2-(2-aminopropylamino)-4-(cyclobutylamino)pyrimidine-5-carboxamide

Step 1: To a stirring solution of alcohol 3.1 (4.16 g, 24 mmol) indichloromethane (30 mL) was added diisopropylethylamine (5.8 mL, 33mmol) followed by methanesulfonyl chloride (2.23 mL, 29 mmol) which wasadded dropwise. After 30 min the reaction was diluted with 1 M HCl andthe two phases separated. The organic layer was then washed with sodiumcarbonate (saturated, aq) and dried over magnesium sulfate.Concentration afforded the desired product 3.2 as a light brown solid(5.07 g, 83%). ¹H NMR (DMSO-d₆, 400 MHz): δ 6.93 (d, 1H), 4.03 (d, 2H),3.72 (m, 1H), 3.15 (s, 3H), 1.37 (s, 9H), 1.06 (d, 3H).

Step 2: Mesylate 3.2 (2.0 g, 7.9 mmol) was dissolved in 20 mL of DMF,treated with sodium azide (2.6 g, 39.5 mmol) and heated to 80° C.overnight. The following day the reaction was cooled to rt, partitionedwith water and ethyl acetate and the two layers separated. The aq phasewas extracted with ethyl acetate and the combined organic phases driedover magnesium sulfate. Concentration afforded the desired azide 3.3which was immediately used for the next step. ¹H NMR (DMSO-d₆, 400 MHz):δ 6.91 (d, 1H), 3.62 (m, 1H), 3.20 (d, 2H), 1.38 (s, 9H), 1.01 (d, 3H).

Step 3: The crude azide from the previous step 3.3 was diluted withmethanol (20 mL), treated with ca. 200 mg of Pd/C (10% by weight, wet)and placed under an atmosphere of hydrogen. After three hours themixture was filtered through celite and concentrated affording aquantitative amount of the desired amine which crystallized uponstanding to a white waxy solid. ¹H NMR (DMSO-d₆, 400 MHz): δ 6.52 (d,1H), 3.38 (m, 1H), 2.42 (m, 2H), 1.36 (s, 9H), 0.96 (d, 3H).

Step 4: Benzotriazolyl ether 1.7, intermediate 3.4,diisopropylethylamine, and 5 mL of 1,4-dioxane were combined and heatedto 120° C. overnight. The reaction mixture was then cooled, diluted withwater and purified by preparative HPLC to give the desired compound. MSfound for C₁₂H₂₀N₆O as (M+H)⁺ 265.2.

Example 6012-((1R,2S)-2-aminocyclohexylamino)-4-(4-bromo-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Compound 601-1 (150 mg, 0.46 mmol) was dissolved in 15 mL DMF. To it wasadded N-bromosuccinimide (NBS, 122 mg, 0.69 mmol). The mixture wasstirred at RT for 12 min. All compound 601-1 had thus been converted tothe corresponding sulfoxide. To it was again added N-bromosuccinimide(122 mg, 0.69 mmol). The mixture was stirred at RT for 1 h. To it wereadded DIEA (0.4 mL, 2.3 mmol) and compound 601-2 (200 mg, 0.92 mmol).The mixture was stirred at 90° C. for 3 h to give products 601-3 and601-4 in ratio of 4:1. The mixture was diluted with ethyl acetate,washed with brine x3, dried, concentrated and subjected to flash columnto isolate compound 601-3 and compound 601-4.

Compound 601-3 was treated with 1:1 TFA and DCM at RT. The mixture wasstirred for overnight at RT. It was concentrated and subjected toreverse phase HPLC to isolate the title compound. MS found forC₁₉H₂₂BrN₉O as (M+H)⁺ 472.3, 474.3. λ=246, 294 nm.

Example 6022-((1R,2S)-2-aminocyclohexylamino)-4-(3-bromo-5-(21-1-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Compound 602-1 (as prepared in Example 601) was treated with 1:1 TFA andDCM at RT. The mixture was stirred for overnight at RT. It wasconcentrated and subjected to reverse phase HPLC to isolate the titlecompound. MS found for C₁₉H₂₂BrN₉O as (M+H)⁺ 472.3, 474.3. λ=253, 272nm.

Example 6032-((1R,2S)-2-aminocyclohexylamino)-4-(4-chloro-3-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Compound 603-1 (150 mg, 0.46 mmol) was dissolved in 15 mL DMF. To it wasadded N-chlorosuccinimide (NCS, 300 mg, 2.2 mmol). The mixture wasstirred at RT for 80 min. To it were added DIEA (0.4 mL, 2.3 mmol) andcompound 603-2 (200 mg, 0.92 mmol). The mixture was stirred at 90° C.for 2 h to give products 603-5 and 603-6 in ratio of 2.6:1. The mixturewas diluted with ethyl acetate, washed with brine x3, dried,concentrated and subjected to flash column to isolate compound 603-5 andcompound 603-6.

Compound 603-5 was treated with TFA at RT. The mixture was stirred for30 m at RT. It was concentrated and subjected to reverse phase HPLC toisolate the title compound. MS found for C₁₉H₂₂ClN₉O as (M+H)⁺ 428.3.λ=246, 291 nm.

Example 6042-((1R,2S)-2-aminocyclohexylamino)-4-(3-chloro-5-(2H-1,2,3-triazol-2-yl)phenylamino)pyrimidine-5-carboxamide

Compound 604-6 (as prepared in Example 603) was treated with TFA at RT.The mixture was stirred for 30 m at RT. It was concentrated andsubjected to reverse phase HPLC to isolate the title compound. MS foundfor C₁₉H₂₂ClN₉O as (M+H)⁺ 428.3. λ=252 nm.

Example 605(S)-4-(4-(1,2,3-thiadiazol-4-yl)phenylamino)-2-(2-aminopropylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 424. UV λ=230, 306 nm. MS found for C₁₆H₁₈N₈OS as (M+H)⁺ 371.3.NMR (CD₃OD): δ 9.23 (s, 1H), 8.54 (s, 1H), 8.15 (m, 2H), 7.81 (m, 2H),3.70 (m, 2H), 3.46 (m, 1H), 1.32 (d, J=6.8 Hz, 3H) ppm.

Example 606(S)-2-(2-aminopropylamino)-4-(4-(thiazol-4-yl)phenylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 424. UV λ=238, 311 nm. MS found for C₁₇H₁₉N₇OS as (M+H)⁺ 370.3.NMR (CD₃OD): δ 9.05 (d, J=2.0 Hz, 1H), 8.51 (s, 1H), 8.00 (d, J=8.0 Hz,2H), 7.89 (s, 1H), 7.70 (d, J=7.2 Hz, 2H), 3.70 (m, 2H), 3.47 (m, 1H),1.30 (d, J=6.0 Hz, 31-1) ppm.

Example 607(S)-2-(2-aminopropylamino)-4-(quinoxalin-6-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 424. UV λ=242, 273 nm. MS found for C₁₆H₁₈N₈O as (M+H)⁺ 339.3.NMR (CD₃OD): δ 8.85 (s, 1H), 8.82 (s, 1H), 8.86 (s, 1H), 8.59 (s, 1H),8.10 (d, J=8.8 Hz, 1H), 7.84 (d, J=−8.4 Hz, 1H), 3.75 (m, 2H), 3.62 (m,1H), 1.38 (d, J=6.0 Hz, 3H) ppm.

Example 608(S)-2-(2-aminopropylamino)-4-(benzo[d]thiazol-5-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 424. UV λ=246, 291 nm. MS found for C₁₅H₁₇N₇OS as (M+H)⁺ 344.3.NMR (CD₃OD): δ 9.34 (s, 1H), 8.72 (s, 1H), 8.56 (s, 1H), 8.11 (d, J=8.8Hz, 1H), 7.52 (d, J=8.8 Hz, 1H), 3.78-3.54 (m, 3H), 1.29 (d, J=6.4 Hz,3H) ppm.

Example 609(S)-2-(2-aminopropylamino)-4-(benzo[d]thiazol-6-ylamino)pyrimidine-5-carboxamide

The title compound was prepared using the same chemistry shown forExample 424. UV λ=241, 297 nm. MS found for C₁₅H₁₇N₇OS as (M+H)⁺ 344.3.NMR (CD₃OD): δ 9.24 (s, 1H), 8.56 (s, 1H), 8.41 (s, 1H), 8.10 (d, J=8.8Hz, 1H), 7.72 (d, J=8.8 Hz, 1H), 3.71-3.44 (m, 3H), 1.23 (d, J=6.4 Hz,3H) ppm.

Example 610

This example illustrates methods for evaluating the compounds of theinvention, along with results obtained for such assays. The in vitro andin vivo human syk activities of the inventive compounds can bedetermined by various procedures known in the art, such as a test fortheir ability to inhibit the activity of human plasma syk. The potentaffinities for human syk inhibition exhibited by the inventive compoundscan be measured by an IC₅₀ value (in nM). The IC₅₀ value is theconcentration (in nM) of the compound required to provide 50% inhibitionof human syk proteolytic activity. The smaller the IC₅₀ value, the moreactive (potent) is a compound for inhibiting syk activity.

An in vitro assay for detecting and measuring inhibition activityagainst syk is as follows:

Inhibition of syk Tyrosine Phosphorylation Activity

Potency of candidate molecules for inhibiting syk tyrosinephosphorylation activity is assessed by measuring the ability of a testcompound to inhibit syk-mediated tyrosine phosphorylation of asyk-specific substrate.

SYK tyrosine phosphorylation activity is measured using the LANCE™Technology developed by Perkin Elmer Life and Analytical Sciences(Boston, Mass.). LANCE™ refers to homogeneous time resolved fluorometryapplications using techniques such as time-resolved fluorescenceresonance energy transfer assay (TR-FRET) (see generally for proceduresin Perkin Elmer Application Note—How to Optimize a Tyrosine Kinase AssayUsing Time Resolved Fluorescence-Based LANCE Detection,wwww.perkinelmer.com/lifesciences). The assay principle involvesdetection of a phosphorylated substrate using energy transfer from aphosphospecific europium-labeled antibody tostreptavidin-allophycocyanin as an acceptor.

To test the ability of candidate molecules to inhibit SYK tyrosinephosphorylation activity, molecules are reconstituted in 30% DMSO andserially diluted 1:3 with the final dilution containing DMSO in theabsence of the candidate molecule. The final DMSO concentration in theassay is 3%. Kinase assays are performed as a two part reaction. Thefirst reaction is a kinase reaction and which comprises of a candidatemolecule, full length active recombinant SYK enzyme (Millipore, Calif.)and biotin-labeled SYK-specific substrate biotin-DEEDYESP-OH. The secondreaction involves termination of the kinase reaction and thesimultaneous addition of the detection reagents—europium-labeledanti-phosphotyrosine reagent (Eu-W1024-PY100, Perkin Elmer, Boston,Mass.) and Streptavidin-Allophycocyanin detection reagent (SA-APC,Prozyme, Calif.). The kinase reaction is performed in a black U-bottom96-well microtitre plate. The final reaction volume is 50 μL andcontains a final concentration of 1 nM active SYK enzyme, 550 nMSYK-substrate, and 100 μM ATP diluted in a buffer containing 50 mM TrispH 7.5, 5 mM MgCl₂, and 1 mM DTT. The reaction is allowed to proceed for1 hour at room temperature. The quench buffer contains 100 mM Tris pH7.5, 300 mM NaCl₂, 20 mM EDTA, 0.02% Brij35, and 0.5% BSA. The detectionreagents are added to the reaction mixture at the followingdilutions-1:500 for Eu-W1024-PY100 and 1:250 for SA-APC. The kinasereaction is terminated by the addition of 50 μL quench buffer containingthe detection reagents. The detection is allowed to proceed for 1 hr atroom temperature. Detection of the phosphorlated substrate in theabsence and presence of inhibitors is measured in the TR-FRETinstrument, Analyst HT (Molecular Probes, Sunnyvale, Calif.) and thecondition for measurements are set up using CriterionHost Release 2.0(Molecular Probes, Sunnyvale, Calif.). The settings used are a follows:excitation 360 nm, emission 665-7.5 nm, beam splitter 350 nm 50/50,flash 100 pulses, delay 60 us, integration 400 us, z-height 2 mm.Inhibition of SYK-tyrosine kinase activity is calculated as the maximumresponse observed in the presence of inhibitor, compared to that in theabsence of inhibitor. IC₅₀s were derived by non-linear regressionanalysis.

Intracellular phospho-flow cytometry was used to test compoundinhibition of Syk activity in intact non-Hodgkin's lymphoma cell linesRamos and SUDHL-6. 10×10⁶ cells in log phase growth were aliqoted; Sykkinase is activated by incubating cells for 10 minutes with 3 μg/mlantibody specific to the B cell receptor. Directly following, cells arefixed in 1% paraformaldehyde for 5 minutes at room temperature, washedin phosphate buffered saline, and then permeablized by incubation for 2hours in ice cold methanol. Cells are again washed in phosphate bufferedsaline, then incubated for 30 minutes with antibody specific forphosphorylated Erk (Y204) and BLNK (Y84), which are indicators of Sykkinase activity, and phosphorylated Syk (Y352), a measure of Src familykinase activity. All antibodies used are purchased from BD Pharmingen(San Jose, Calif.). After incubation with antibodies, cells are againwashed and subjected to flow cytometry. Representative data detailinginhibition of B cell receptor singaling by compounds are shown in Table1 as IC₅₀ ranges.

The anti-proliferative effects of compounds on non-Hodgkin's lymphoma Bcell lines SUDHL-4, SUDHL-6, and Toledo was also assessed. SUDHL-4 andSUDHL-6 require B cell receptor signaling for growth and survival, whilethe Toledo cell line (serving here as a negative control) does not.Cells were aliquoted into each well of a 96-well plate and incubatedwith increasing concentrations of compound for 72 hours, after whichcell survival and proliferation was determined using the MTT assay(Chemicon International, Inc., Temecula, Calif.) following protocolssupplied by the manufacturer. Data are detailed in the Tables andFigures herein as IC₅₀ values plus or minus standard deviations from 5or 6 independent experiments.

Induction of apoptosis in non-Hodgkin's lymphoma B cell lines SUDHL-4,SUDHL-6, and Toledo was assessed by measuring the apoptotis markerCaspase 3. Cells were incubated with 1, 3, or 10 μM compound for 24, 48,and 72 hours. At the conclusion of each time point, cells were processedfor flow cytometry analysis using the Monoclonal Rabbit Anti-ActiveCaspase-3 Antibody Kit and related protocols (BD Pharmingen). Data fromtwo independent experiments are presented in Tables 7A and 7B,representing the percent of total cells undergoing apoptosis followingincubation with compounds under the indicated conditions.

Syk activity is not only required for B cell signaling, proliferation,and survival, as shown, but is also critical for cellular activationupon cross-linking of the B cell receptor. B cell activation leads toincreased cell surface expression of several proteins involved in cellsignaling, antigen presentation, and adhesion. Among these, CD80, CD86,and CD69 are commonly measured to determine B cell activation status.Therefore, primary mouse B cells isolated from spleen were aliquoted andincubated with increasing concentrations of compound (0.05 to 2 μM) inthe presence of goat anti-mouse IgD (eBiosciences, Inc., San Diego,Calif.) for 20 hours to cross-link the B cell receptor. Following, cellswere washed and incubated for 30 minutes on ice with antibodies specificfor the CD80, CD86, and CD69 B cell activation markers. B cells wereidentified from the pooled population by staining with the B cell markerCD45RO. All antibodies were purchased from BD Pharmingen. Table 8depicts the IC₅₀ range in which these compounds inhibited B cellreceptor induced activation of mouse primary B cells

In the table below, activity in the Syk and/or Jak assays is provided asfollows: +++++=IC₅₀<0.0010 μM; ++++=0.0010 μM<IC₅₀<0.010 μM, +++=0.010μM<IC₅₀<0.10 μM, ++=0.10 μM<IC₅₀<1 μM, +=IC₅₀>1 μM.

TABLE 6a Example UV MW MH+ Syk IC50  1 240, 297 452.6 453.2 ++  2 236,312 459.6 460.1 ++  3 239, 304 392.5 393.2 ++  4 231, 311 410.5 411.2+++  5 238, 337 403.5 404.2 +++  6 239, 311 403.5 404.2 +++  7 241, 330403.5 404.2 +++  8 243, 308 404.5 405.2 +++  9 239, 313 409.5 410.2 +++410.4  10 233, 308 410.5 411.2 +++  11 240, 311 403.5 404.2 +++  12 245404.5 405.2 +++ 405.3  13 240, 312 403.5 404.2 +++  14 240, 318 409.5410.2 +++  15 243, 332 409.5 410.2 +++  16 240, 302 392.5 393.2 +++393.3 393.4  17 248 403.5 404.2 ++++  18 243, 294 411.5 412.2 +++ 412.3 19 242, 307 404.5 405.3 +++  20 244, 293 425.5 426.3 ++  21 239, 327404.5 405.4 +++  22 235, 319 404.5 405.4 +++  23 239, 334 403.5 404.4+++  24 242, 300 393.5 394.4 +++  25 241, 300 406.5 407.4 +++  26 241,296 397.4 398.4 ++  27 246, 292 392.5 393.2 +++ 393.5  28 246, 287 409.5410.5 ++  29 239, 309 429.5 430.4 +++  30 241, 236 460.5 461.4 ++  31247 392.5 393.4 ++++  32 410.5 411.3 ++ 411.5  33 427.5 428.4 +++  34410.5 411.3 +++  35 424.5 425.3 ++  36 201.6, 240.4, 410.5 411.09 ++289.0  37 202.8, 245.2 409.5 410.2, 411.2 ++++  38 240.4, 312.8 427.5428.5, 429.4 +++  39 206.3, 242.8, 409.5 410.2, 411.4 ++++ 291.4  40201.6, 244.0, 409.5 410.5, 411.6 ++++ 277.1  41 242.8, 292.6 403.5404.2, 405.3 +++  42 244.0, 289.0 404.5 405.3, 406.5 +++  43 246.3 404.5405.2, 406.2 +++  44 242.8 406.5 407.2, 408.2 +++  45 247.5 433.5 434.3,435.3 +++  46 240.4, 306.8 433.5 434.3, 435.3 +++  47 240.4, 296.1 425.5426.3, 427.3 +++  48 418.5 419.5 (M + 1) +++  49 438.6 439.6 (M + 1) ++ 50 393.5 MS: 394.2 (M + H) +++  51 393.5 MS: 394.3 (M + H) +++  52394.4 MS: 395.28 (M + H) ++  53 393.5 MS: 394.28 (M + H) +++  56 408.5MS: 409.28 (M + H) +++  57 423.5 MS: 424.37 (M + H) +++  58 408.5 MS:409.32 (M + H) +++  59 392.5 MS: 393.0 (M + H) ++++  60 408.5 MS: 410(M + H) +++  61 423.5 MS: 424.5 (M + H) +++  62 406.5 MS: 407.5 (M + H)+++  63 409.5 MS: 410.5 (M + H) +++  64 420.5 MS: 421.5 (M + H) ++  65424.5 MS: 425.4 (M + H) ++++  66 394.4 MS: 395.5 (M + H) ++++  67 408.5MS: 409.5 (M + H) +++  68 408.5 MS: 4095 (M + H) +++  69 408.5 MS: 409.5(M + H) +++  70 391.5 MS: 392.5 (M + H) ++++  71 391.5 MS: 392.4 (M + H)+++  72 406.5 MS: 407.5 (M + H) +++  73 395.5 MS: 396.6 (M + H) +++  74411.5 MS: 412.5 (M + H) +++  75 394.4 MS: 395.3 (M + H) +++  76 394.4MS: 395.2 (M + H) ++++  78 394.4 MS: 395.2 (M + H) +++  79 243, 302478.5 479.4 ++  80 246, 301 594.5 595.4 +  81 247 477.6 478.4 +  82 241392.5 393.4 ++++  83 241, 319 424.5 425.3 +++  84 246, 293 406.5 407.3++  85 243, 285 406.5 407.3 ++  86 244 393.5 394.3 ++++  87 250 393.5394 ++++ 394 (M + H) 394.3 394.8 (M + H) ES (+) MS ES (+) MS [M + 1] =394  88 242, 300 423.5 424.3 +++  89 242, 301 422.5 423.3 +++  90 242,300 436.5 437.3 +++  91 249, 301 489.6 490.3 +  92 247 411.4 412.3 +++ 93 259 418.5 419.4 +  94 252 418.5 419.4 ++  95 244, 288 418.5 419.4 ++ 96 245, 296 433.5 434.4 +++  98 249 422.5 423.4 +++ 101a 250 394.4395.3 +++ 101b 250 408.5 409.3 ++++ 102 246, 304 407.5 408.3 +++ 103 249442.5 443.3 +++ 104a 240, 295 442.5 443.4 +++ 104b 240, 295 442.5 443.4+++ 105a 234, 303 443.5 444.4 +++ 105b 234, 303 443.5 444.4 +++ 106 240,292 442.5 443.3 +++ 107 243, 290 459.6 460.3 ++ 108 245 486.6 487.5 ++109 263 453.6 454.3 +++ 110 249 453.6 454.3 +++ 111 240, 311 406.5 407.3+++ 113 239, 314 406.5 407.3 ++++ 114 239, 310 393.5 394.3 +++ 115 250411.4 412.3 +++ 116 244, 295 423.5 424.3 ++++ 117 249 423.5 424.3 ++++118 243, 281 407.5 408.3 ++++ 119 248 407.5 MW = 407.47, ++++ M + H =408.8 120 249 404.5 405.3 +++ 405.4 121 231, 314 404.5 405.3 +++ 122 249406.5 407.3 ++++ 123 247 406.5 407.3 +++ 124 247 442.5 443.3 +++ 125245, 303 442.5 443.3 +++ 127 241, 298 383.5 384.1 +++ 384.2 128 246, 295367.4 368.1 +++ 129 246, 295 397.5 398.1 +++ 130 239, 309 365.4 366.2++++ 132 241, 283 377.5 378.1 ++++ 378.2 (M + 1) 133 243, 294 366.4367.2 +++ 134 246, 292 383.5 384.2 +++ 135 250 366.4 367.1 +++ 136 240,294 384.4 385.1 +++ 137 242 378.4 379.2 +++ 138 243 384.5 385.1 ++ 139391.5 392.2 (M + 1) +++ 140 377.5 378.34 +++ 142 377.5 378.34 (M + 1) ++143 476.6 477.40 (M + 1) +++ 144 406.4 407.28 (M + 1) ++ 146 381.4382.35 (M + 1) +++ 147 439.5 440.38 (M + 1) ++ 148 397.4 398.31 (M + 1)+++ 149 409.5 410.36 (M + 1) +++ 150 402.5 403.32 (M + 1) +++ 151 395.5396.35 (M + 1) ++++ 152 393.5 394.14 (M + 1) ++++ 153 411.5 412.5(M + 1) +++ 154 397.4 398.5 (M + 1) ++ 155 407.5 408.6 (M + 1) +++ 156468.6 469.4 + 157 377.5 378.2 (M + 1) ++++ 378.5 (M + 1) 378.6 (M + 1)158 380.5 381.17 (M + 1) ++++ 159 380.5 381.17 (M + 1) ++++ 160 380.5381.17 (M + 1) +++ 161 462.6 463.32 (M + 1) +++ 162 397.5 398.2 (M + 1)+++ 163 379.5 380.4 (M + 1) +++ 165 432.5 433.2 (M + 1) ++++ 166 411.5412.2 (M + 1) ++ 167 245 495.6 496.3 +++ 168 249 499.6 500.3 +++ 169 244495.6 496 +++ 496.3 170 246 499.6 500.3 +++ 171 247 487.6 488 +++ 488.3172 246 487.6 488.3 +++ 173 238, 309 487.6 488.3 +++ 174 241, 314 487.6488.3 +++ 175 246 408.5 409 +++ 409.4 176 241, 290 410.5 411.4 ++ 177239, 309 408.5 409.3 +++ 409.4 178 243, 296 410.5 411.3 ++ 179 232, 306466.6 467.3 ++ 180 238, 290 418.5 419.3 ++ 181 241, 291 426.5 427.3 +++183 244, 297 439.5 440.3 ++ 185 243.8, 290.0 456.5 457.2 ++ 186 400.4401.2, ++ racemate 187 242.6, 290.0 456.5 457.2, ++ racemate 188 242.6,290.0 400.4 401.2, +++ racemate 189 241.2, 292.1 330.4 331.4 ++ 190237.6, 313.1 329.4 330.3 ++ 191 240.0, 303.2 392.5 393.4 +++ 192 244.9,326.1 346.5 347.3 ++++ 193 247.3, 325.6 346.5 347.3 +++ 194 240.0, 314.3391.5 392.4 +++ 195 244.0, 316.4 446.6 447.3, 448.4 +++ 196 240.4, 304.5447.5 448.1, 449.2 ++ 197 239.3, 308.7 402.5 403.4 +++ 198 243.6, 286.5393.5 394.4, 395.5 ++++ 199 245.2, 293.8 393.5 394.4, 395.4 ++ 200 239.9393.5 394.5 ++ 202 243.6, 287.1 407.5 408.4 +++ 203 241.4, 288.8 412.5413.1, 414.2 ++ 204 240.5, 287.8 398.5 399.2, 400.2 +++ 205 238.1, 292.6370.5 371.2, 372.3 +++ 206 236.9, 286.6 386.5 387.1, 388.2 +++ 207240.4, 292.6 418.5 419.3, 420.3 ++ 209 376.5 377.1, 378.5 +++ 210 227.5,319.9 406.5 407.2, 408.5 ++++ 211a 212.2, 244.0, 394.5 395.1, 396.3 ++++306.8 211b 223.9, 318.8 419.5 420.2, 421.4 ++++ 212 219.2, 235.7, 433.5434.3, 435.3 +++ 318.8 213 218.0, 314.0 447.5 448.2, 449.4 ++++ 214223.9, 293.8 410.9 411.2, 413.1 ++ 215 455.4 455.1, 457.1 ++++ 216 381.4382.3 +++ 217 220.4, 315.2 489.6 490.4, 491.4 ++++ 218 219.2, 241.6,379.5 380.4 ++++ 336.7 219 395.5 396.3 ++++ 219 216.7, 239.9, 365.4366.3, 367.4 +++++ 330.3 220 210.6, 243.0, 380.5 381.5 ++++ 329.1 221238.1 381.4 382.4 ++ 222 234.5, 298.5, 384.5 385.3 +++ 315.2 223 203.4,245.4 378.4 379.3 +++ 224 205.1, 242.8, 383.5 384.3 +++ 290.2 225 202.8,239.3 380.5 381.4 ++ 226 220.4, 239.3, 379.5 380.4 ++++ 336.7 227 241.6,293.8 441.5 442.5 ++ 228 222.8, 242.8 393.5 394.4 +++ 229 222.8, 241.6405.5 406.5 ++++ 230 203.9, 236.9, 367.4 368.4 +++ 294.9 231 220.2,242.6 393.5 394.4 ++++ 232 210, 243 462.6 MW = 462.5; +++ M + 1 = 463.3233 248 432.5 MW = 432.5, +++ M + 1 = 433.4 234 245 492.6 MW = 492.6, ++M + 1 = 493.4 235 241 432.5 MW = 432.5, +++ M + 1 = 433.3 236 245, 276444.5 445 ++++ MW = 444.5, M + 1 = 445.3 237 244, 288 432.5 MW = 432.5,+++ M + 1 = 433.3 238 245 420.5 MW = 420.5; ++++ M + 1 = 421.3 239 242420.5 MW = 420.5; +++ M + 1 = 421.3 240 246 462.6 MW = 462.5; +++ M + 1= 463.3 242 245 438.5 MW = 438.5; +++ M + 1 = 439.3 243 244, 288 462.6463.3 +++ 245 211, 243 460.5 MW = 460.5; ++++ M + 1 = 461.2 246 242, 316460.5 MW = 460.5; +++ M + 1 = 461.2 247 216, 238, 303 473.6 MW = 473.6,+++ M + 1 = 474.4 248 252 485.6 MW = 485.6; +++ M + 1 = 486.4 249 247485.6 MW = 485.6; +++ M + 1 = 486.4 250 246 471.6 MW = 471.6; +++ M + 1= 472.4 251 248, 282 485.6 MW = 485.6; +++ M + 1 = 486.4 252 245 485.6MW = 485.6; +++ M + 1 = 486.4 253 239, 274, 296 446.5 MW = 446.5; ++++M + 1 = 447.3 254 240, 285 480.6 MW = 480.6; + M + 1 = 481.4 255 246480.6 MW = 480.6; ++++ M + 1 = 481.4 256 250 480.6 MW = 480.6; +++ M + 1= 481.4 257 235, 308 480.6 481.4 +++ 259 239, 313 480.6 481.4 +++ 260239, 302 453.6 454.4 ++ 261 240, 301 453.6 454.4 ++ 262 241, 302 453.6454.4 ++ 264 212, 236, 322 416.5 417.4 +++ 265 210, 244, 289 416.5 417.3+++ 266 238, 296 429.5 430.3 +++ 267 250 393.5 394.3 ++ 268 250 393.5394.3 +++ 269 252 393.5 394.3 ++ 270 252 393.5 394.3 + 272 252 393.5394.3 + 273 251 379.4 380.3 + 274 259 461.5 MW = 461.4, +++ M + 1 =462.2 275 239, 296 407.5 MW = 407.47, +++ M + 1 = 408.4 276 238, 296406.5 MW = 406.48, ++ M + 1 = 407.5 277 245, 295 461.5 MW = 461.4, +++M + 1 = 462.4 278 245, 295 461.5 MW = 461.4, ++ M + 1 = 462.4 279 244352.4 353.2 ++++ 280 236, 310 352.4 353.2 +++ 281 242, 289 354.5 355.3+++ 282 240, 290 368.5 369.3 +++ 283 250 407.5 408.4 +++ 284 391.4 392.4+++ 285 242, 305 411.4 MW = 411.44, +++ M + 1 = 412.2 292 376.5 377.1++++ 377.1 (M + 1) 377.3 (M + 1) 3771., 378.2 294 245.4 379.5 380.4 ++295 249 379.4 380.4 ++ 296 240, 287 326.4 327.4 ++ 297 217, 239, 290340.4 341.4 +++ 298 249 395.4 396.3 +++ 299 250 395.4 396.3 ++ 300 246394.4 395.3 +++ 301 249 406.5 407.3 ++ 302 240, 288 342.4 343.3 +++ 303240, 290 356.4 357.3 ++++ 304 240, 283 358.4 359.3 +++ 305 241, 287342.4 343.3 ++ 306 239, 292 376.4 377.3 ++ 308 240, 292 358.4 359.2 +++310a 243, 294 356.4 357.3 ++ 310b 243, 290 356.4 357.3 +++ 311 249 409.5410.3 +++ 312a 247, 295 370.5 371.3 ++ 312b 242, 289 370.5 371.3 ++ 314240 356.4 357.3 ++ 315 246, 303 382.4 383.1 +++ 316 245, 302 340.3 341.1++ 317 249, 298 402.4 403.1 ++ 318 241, 283 393.5 394.1 +++ 394.2(M + 1) 319 243, 303 399.5 399.1 +++ 320 245 381.4 382.1 +++ 321 247,301 382.4 383.2 +++ 322 249, 285 420.5 421.1 + 323 250, 300 420.5 421.1++ 324 243, 296 468.6 469.2 ++ 325 244, 297 382.4 383.2 +++ 327 245, 299340.3 341.2 ++ 328 245, 296 413.5 414.2 ++ 330 244, 303 357.4 358.2 ++331 249, 292 357.4 358.2 ++ 332 247, 295 399.5 400.2 +++ 333 250 340.3341.1 ++ 334 252 382.4 383.1 +++ 335 224 339.4 340.1 ++ 336 243, 310400.4 401.1 ++ 337 243, 312 358.4 359.1 ++ 338 243 394.4 395.2 +++ 339242 352.4 353.2 ++ 340 241, 308 408.5 409.2 ++ 341 240, 314 425.5 426.2+++ 342 233, 311 426.5 427.2 +++ 343 240 400.5 406.1 ++ 344 244, 314419.5 420.2 ++ 345 244, 294 427.5 428.3 +++ 346 240, 310 445.5 446.3 ++347 246 427.5 428.3 ++ 349 247, 292 411.5 412.3 ++ 350 241, 314 419.5420.3 ++ 351 250 419.5 420.3 ++ 352 243, 284 395.5 MS: 396.44 (M + H)+++ 353 250 408.5 409.3 ++ 354 367.4 MS: 368.17 (M + H) ++ 355 367.4 MS:368.29 (M + H) ++ 356 450.5 451.35 (M + 1) +++ 357 251 409.5 410.3 ++358 250 381.4 382.3 ++ 359 251 367.4 368.3 ++ 360 247 353.3 354.3 + 361240, 313 424.5 425.3 +++ 362 244, 298 426.5 427.3 ++ 364 251, 302 490.6491.3 ++ 372 207.5, 244.0, 396.5 397.4 ++ 325.9 374 388.4 Turbo Spray ++Mass Spec [M + 1] = 388 375 407.3 Turbo Spray ++ MS [M + 1] = 407 376372.4 Turbo Spray ++ MS [M + 1] = 373 378 396.4 Turbo Spray ++ MS [M +1] = 397 379 353.4 Turbo Spray ++ MS [M + 1] = 354 380 358.4 Turbo Spray++ MS [M = 1] = 359 381 342.4 Turbo Spray ++ MS [M + 1] = 343 383 360.4Turbo Spray ++ [M + 1] = 361 385 328.4 Turbo Spray ++ MS [M + 1] = 329386 314.3 ES (+) MS [M + H] = 315 ++ Turbo Spray MS [M + 1] = 315 387330.3 Turbo Spray ++ MS [M + 1] = 331 389 422.4 Turbo Spray ++ Mass Spec[M + 1] = 423 390 392.4 Turbo Spray ++ MS [M + 1] = 393 391 430.4 TurboSpray ++ [M + 1] = 431 392 441.3 Turbo ++ Spray = 442 393 344.4 TurboSpray ++ MS [M + 1] = 345 394 344.4 Turbo Spray ++ MS [M + 1] = 345 396330.3 Turbo Spray ++ MS [M + 1] = 331 397 395.2 Turbo Spray ++ MS [M] =395 398 346.3 Turbo Spray ++ MS [M + 1] = 347 399 404.5 Turbo Spray ++MS [M + 1] = 405 400 424.9 Turbo Spray ++ MS [M + 1] = 425 401 408.4Turbo Spray ++ MS [M + 1] = 409 402 404.5 Turbo Spray ++ MS [M + 1] =405 404 391.4 Turbo Spray ++ MS [M + 1] = 392 405 420.5 Turbo Spray ++MS [M + 1] = 421 406 406.4 Turbo Spray +++ MS [M + 1] = 407 407 406.4Turbo Spray ++ MS [M + 1] = 407 408 248, 294 385.5 386.4 + 409 240, 283365.4 366.2 +++ 410 243, 302 385.5 386.4 ++ 411 246, 293 371.4 372.4 ++412 216.9, 244.0 395.5 396.3 +++ 413 213.3, 244.0, 396.5 397.4 +++ 333.1414 241.6, 318.8 382.4 383.4, 384.3 ++ 416 240, 292 300.4 301.3 +++ 417249 339.4 340.3 +++ 418 251 353.4 354.3 ++ 419 252 367.4 368.3 ++ 420249 367.4 368.3 ++ 422 250 353.4 354.3 ++ 423 254 394.4 395.3 + 424 250353.4 354.3 +++ 425 247 364.4 365.4 +++ 426 243, 289 300.4 301.4 +++ 427243, 289 314.4 315.4 +++ 428 250 353.4 354.3 +++ 440 219.2, 239.3, 369.4370.4 ++++ 331.9 441 220.4, 325.9 367.5 368.4 ++++ 442 220.4, 239.9,397.5 398.4 ++++ 331.6 443 219.2, 239.3, 383.5 384.3 ++++ 331.9 444215.7, 238.1, 369.4 370.3 ++++ 327.1 445 214.9, 241.7, 370.4 371.3 ++++324.1 447 203.9, 244.0, 395.5 396.4 ++++ 303.3 448 208.6, 240.4, 384.4385.4 +++ 283.1, 324.7 449 221.6, 336.7 409.5 410.4 +++ 450 222.8, 336.7409.5 410.4 +++ 451 221.6, 335.5 409.5 410.4 +++ 452 219.2, 239.3, 381.5382.4, 383.5 +++ 327.1 454 439.9 Turbo Spray +++ MS [M + 1] = 440 455396.4 Turbo Spray +++ MS [M + 1] = 396 456 405.5 Turbo Spray +++ MS [M +1] = 406 457 359.4 ES (+) MS +++ [M + 1] = 360 458 426.5 Turbo Spray +++MS [M + 1] = 427 459 460.9 Turbo Spray +++ MS [M + 1] = 447 460 435.5Turbo Spray ++ MS [M + 1] = 436 462 423.5 Turbo Spray ++ MS [M + 1] =424 463 462.5 Turbo Spray ++ MS [M + 1] = 463 465 246 367.4 MW = 367.4,+++ M + 1 = 368.3 467 238, 284 314.4 415.4 +++ 468 240, 290 328.4 329.4+++ 469 248 367.4 368.4 ++++ 470 248 378.4 379.4 +++ 471 246 366.4 367.4+++ 473 250 379.4 380.4 +++ 478 245 421.4 422.3 +++ 479 240 340.4 341.5+++ 481 219.2, 240.4, 367.5 368.4 ++++ 331.9 482 220.2, 241.4, 365.4366.4 ++++ 330.6 483 219.2, 240.4, 353.4 354.4 ++++ 333.1 484 219.3,239.4, 397.5 398.4 ++++ 326.7 485 216.7, 238.7, 355.4 356.4 ++++ 327.2499 224.9, 316.2 409.5 410.2, 411.3 +++ 500 223.9 383.5 384.4 +++ 501203.9, 241.6 384.4 385.3, 386.4 +++ 502 220.4, 333.1 369.4 370.4 +++ 503209.8, 240.4, 384.4 385.4 +++ 283.1, 325.9 507 220.4, 240.4 383.5 384.4+++ 508 218.6, 242.3 431.5 432.4 +++ 509 219.8, 241.7 381.5 382.4 +++510 240.3, 290.0 344.4 345.4, 346.5 +++ 511 218 383.5 384.4 +++ 512211.3, 242.3, 432.5 433.4 +++ 330.9 513 202.8, 246.3 392.5 393.4 +++ 514221.6, 315.2 396.5 397.0, 398.4 +++ 515 203.4, 246.6 454.5 455.4 +++ 516222.8, 316.4 423.5 424.4, 425.1 +++ 517 208.8, 241.7, 370.4 371.5 +++316.7 518 240.5, 288.4 328.4 329.5 +++ 519 241.1, 286.5 314.4 315.4 +++520 206.3, 240.4, 356.4 357.3 +++ 311.6 521 203.8, 235.5, 376.4 377.30,378.42 +++ 290.0 522 218.0, 311.6 479.5 480.4 +++ 523 239.3, 289.0 330.3331.1, 332.1 +++ 524 241.1, 287.7 354.5 355.5 +++ 525 241.7, 285.9 328.4329.4 +++ 526 219.2, 282.8 366.4 367.1, 368.2 +++ 367.4 527 241.7, 286.5326.4 327.4 +++ 530 241.4, 287.7 340.4 341.4 +++ 531 242.8 393.5 394.4+++ 532 240.4, 290.2 399.5 400.2, 401.5 +++ 533 204.6, 251.5 393.5 394.4+++ 535 242.3, 285.4 383.4 384.3 +++ 536 239.3, 285.4 344.4 345.1, 356.1+++ 537 202.8, 238.1, 370.4 371.3 +++ 308.0 538 208.6, 240.4, 384.4385.3 +++ 319.9 539 203.9, 249.9 367.4 368.4 +++ 540 202.1, 246.0 406.5407.4 +++ 543 215.7, 240.4, 364.8 365.2, 367.1 +++ 289.0 546 239.9,284.7 342.4 343.5 +++ 547 238.1, 299.7 370.4 371.4 +++ 548 239.7 346.4347.23, 348.32 +++ 549 239.3, 286.6 344.4 345.2, 346.3 ++ 550 215.5,241.7, 433.5 434.5 ++ 330.9 551 249.9 383.4 384.3, 385.3 ++ 552 247.2382.4 383.3 ++ 553 243.6 394.4 395.3, 396.4 ++ 554 238.1, 286.6 316.4317.25, 318.83 ++ 556 239.3 344.4 345.2, 346.3 ++ 557 240.5, 286.5 344.4345.4 ++ 558 220.4, 239.3 339.4 340.4 ++ 560 239.3 360.4 361.2, 362.2 ++561 238.1, 281.9 382.4 383.5, 384.5 ++ 562 250.3 411.5 412.4 ++ 563208.6, 240.4, 384.4 385.3 ++ 311.6 564 238.1, 292.6 360.4 361.2, 362.3++ 565 232.6, 287.8 373.4 374.3 +++ 566 239.3, 287.8 356.5 357.4 ++ 567204.6, 250.9 379.4 380.4 ++ 569 242.8 384.4 385.2, 386.4 ++ 570 238.1,290.2 344.4 345.1, 346.2 ++ 581 243.6, 292.1 373.4 374.3 ++ 582 239.3,285.4 342.4 343.4 ++ 583 235.7, 285.4 376.4 377.1, 378.2 ++ 584 240.4,287.8 383.5 384.14, 385.06 ++ 585 235.7, 315.2 374.4 375.2 ++ 586 230.1,311.2 394.4 395.4, 396.3 ++ 587 245.4 368.4 369.2, 370.1 ++ 588 373.5374.3, 375.3 ++ 589 202.8, 239.9, 390.4 391.4 ++ 282.2 590 238.1, 293.8415.5 416.2, 417.4 ++ 591 239.3, 292.7 374.4 375.3 ++ 592 240.4 356.5357.4 ++ 593 241.7, 288.4 408.5 409.4 ++ 594 203.9, 235.7, 357.4 358.3++ 292.6 595 239.3 358.4 359.2, 360.4 ++ 598 248.7 395.5 396.4 ++Inhibition of GPVI-Mediated Platelet Function In Vitro

The ability for candidate molecules to inhibit syk-mediated plateletfunctions are tested by measuring the inhibition the GPVI-specificagonist Convulxin-induced human platelet calcium-mobilization oraggregation. Calcium-mobilization is assessed in human washed plateletsin a 96-well microtiter format. Aggregation is assessed in a 96-wellmicrotiter assay (see generally the procedures in Jantzen, H. M. et al.(1999) Thromb. Hemost. 81:111-117) or standard cuvette lighttransmittance aggregometry using human platelet-rich plasma (PRP).

Inhibition of Convulxin-Mediated Platelet Calcium-Mobilization In Vitro

Inhibition of Convulxin-induced calcium-mobilization was determined inhuman washed platelets using the FLIRP Calcium 3 Assay Kit (MolecularDevices, Sunnyvale, Calif.). For preparation of washed platelets, humanvenous blood is collected from healthy, drug-free volunteers into ACD(85 mM sodium citrate, 111 mM glucose, 71.4 mM citric acid) containingPGI₂ (1.25 ml ACD containing 0.2 μM PGI₂ final; PGI₂ was from Sigma, St.Louis, Mo.). Platelet-rich plasma (PRP) is prepared by centrifugation at160×g for 20 minutes at room temperature. Washed platelets are preparedby centrifuging PRP for 10 minutes at 730 g and resuspending theplatelet pellet in CGS (13 mM sodium citrate, 30 mM glucose, 120 mMNaCl; 2 ml CGS/10 ml original blood volume). After incubation at 37° C.for 15 minutes, the platelets are collected by centrifugation at 730 gfor 10 minutes and resuspended at a concentration of 3×10⁸ platelets/mlin Hepes-Tyrode's buffer (10 mM Hepes, 138 mM NaCl, 5.5 mM glucose, 2.9mM KCl, 12 mM NaHCO₃, pH 7.4). This platelet suspension is kept >45minutes at room temperature before use in calcium mobilization assays.

For 96-well plate Calcium-mobilization experiments, equal volumes of3×10⁸ washed platelets/ml were incubated with equal volumes of Calcium-3Assay Reagent A resuspended in 1× Hank's Balanced Salt Solution, pH 7.4,20 mM Hepes buffer. The total reaction volume of 0.2 ml/well includes1.5×10⁸/ml washed platelet/Calcium-3 Assay reagent A mix, 10 μMEptifibatide (Millennium Pharmaceuticals Inc, Cambridge, Mass.), serialdilutions (1:3) of test compounds in 0.75% DMSO. DMSO alone is added to1 well of each 8 set to allow for a maximal calcium-mobilizationreading. After 20 minutes preincubation at room temperature the 96-wellmicroplate reader is loaded into the FlexStation (Molecular Devices,Sunnyvale, Calif.). The FlexStation experimental conditions formeasuring Calcium mobilization are set up using SOFTMax Pro. Thesettings used are detailed below. Fluorescence parameters-assay mode:flex, excitation 485 nM, 525 nM with a cut-off of 515 nM; Parameters—PMTsensitivity-6, pipette height 230 μl, read time 2 minutes and 40seconds, read intervals 2 seconds, temperature-23-25° C. After 18seconds of baseline reading, calcium-mobilization is initiated by theaddition of Convulxin to a final concentration of 125 ng/ml. Inhibitionof calcium-mobilization was calculated as the maximum response observedin the presence of inhibitor, compared to that in the absence ofinhibitor. IC₅₀s were derived by non-linear regression analysis.

Inhibition of Convulxin-Mediated platelet Aggregation In Vitro

For preparation of human platelet-rich plasma for aggregation assays,human venous blood was collected from healthy, drug-free volunteers into0.38% sodium citrate (0.013 M, pH 7.0 final). Platelet-rich plasma (PRP)is prepared by centrifugation of whole blood at 160×g for 20 minutes atroom temperature. The PRP layer is removed, transferred to a new tube,and the platelet count is adjusted, if advantageous, to achieve aplatelet concentration of ˜3×10⁸ platelets/ml using platelet-poor plasma(PPP). PPP is prepared by centrifugation of the remaining blood sample(after removal of PRP) for 20 minutes at 800×g. This preparation of PRPcan subsequently be used for aggregation assays in either a 96-wellplate or standard cuvette aggregometry.

Inhibition of Convulxin-induced aggregation is determined in 96-wellflat-bottom microtiter plates using a microtiter plate shaker and platereader similar to the procedure described by Frantantoni et al., Am. J.Clin. Pathol. 94, 613 (1990). All steps are performed at roomtemperature. For 96-well plate aggregation using platelet-rich plasma(PRP), the total reaction volume of 0.2 ml/well includes 190 μl of PRP(˜3×10⁸ platelets/ml, see above), and 5 μl of either serial dilution oftest compounds in 30% DMSO or buffer (for control wells). After 20minutes preincubation at room temperature 5 μl of 320 ng/ml Convulxinagonist solution is added to each well to give a final concentration of8 ng/ml Convulxin. The plates are then agitated for 5 min on amicrotiter plate shaker and the 5 minute reading is obtained in themicrotitre plate reader (Softmax, Molecular Devices, Menlo Park,Calif.). Aggregation is calculated from the decrease of OD at 650 nm att=5 minutes. IC₅₀s were derived by non-linear regression analysis.

Inhibition of Convulxin-induced aggregation was also determined forcuvette light transmittance aggregation assays, serial dilutions (1:2)of test compounds were prepared in 30% DMSO in a 96 well V-bottom plate(final DMSO concentration in the cuvette was 0.3%). The test compound (5μl of serial dilutions in DMSO) was preincubated with PRP for 20 minutesprior to initiation of aggregation reactions, which is performed in aChronoLog aggregometer by addition of agonist (125-250 ng/ml Convulxin)to 495 μL of PRP at 37° C. The aggregation reaction is recorded for 4min, and maximum extent of aggregation is determined by the differencein extent of aggregation at baseline, compared to the maximumaggregation that occurs during the 4 minute period of the assay.Inhibition of aggregation was calculated as the maximum aggregationobserved in the presence of inhibitor, compared to that in the absenceof inhibitor. IC₅₀s were derived by non-linear regression analysis.

Examples of compounds and their syk and PRP IC₅₀ values are given intables 2-5.

Calcium Flux Assay in Ramos Cells Induced by Bcr Cross-linking

Ramos cells (2G6.4C10, Burkitt's lymphoma, ATCC Item Number: CRL-1923)are sub-cultured at 5×10⁵ cells/ml in fresh medium 3 or 4 days ahead ofexperiments. Cells are harvest and re-suspend in fresh medium at 8×10⁶cells/ml before dye-loading. An equal volume of Calcium 3 loading dye(Molecular Device) is added and mixed into cell suspension. Loadingcells are dispensed in a 96 well plate and incubated 30 min. Compoundsare then added in the dye-loaded cells and incubated for another 30 min.Spin cell down at 1000 rpm for 3 min before fluorescence measurement inFlexStation. BCR stimulation is carried by the addition of 5 μg/mlantibody (AffiniPure F(ab′)₂ fragment Donkey anti-human IgM, JacksonImmunoResearch Laboraotries).

Calcium Flux Assay in Jurkat Cells Induced by Tcr Cross-linking

The protocol is very similar to B cell calcium flux as described in theprevious section. The only differences are that T cells (clone E6-1,Acute T cell Leukemia, ATCC Item Number: Tib-152) and anti-human CD3(Functional Grade Purified anti-human CD3, clone OKT3, eBioscience, No.16-0037) replaced B cells and anti-human IgM. Cell density is kept thesame but antibody is used at a concentration of 100 ng/ml.

IL-2 Secretion in Jurkat Cells Induced by TCR Cross-linking

Jurkat cell propagation and compound incubation procedures are the sameas described in Jurkat calcium flux assay in the previous section.Antibody (anti CD3, OKT3) is coated onto a fresh plate (without cells)at 100 ng/well. Cells are suspended at 8×10⁶ cells/ml and incubated withcompounds for 30 min in a separate plate. At the end of incubation,cells are transferred to the antibody-coated plate and incubated for 16hours. 100 μl of cell medium after incubation is used for IL-2measurement after incubation. IL-2 level is determined using an IL-2ELISA kit (Human IL-2 ELISA kit II, BD Bioscience, No. 550611).

Example 611 Millipore Upstate KinaseProfiler™ Screening

This assay is a direct measurement of the effect of compound on thecatalytic activity of JAK3. Purified human JAK3 (GenBank AF513860)sequence (residue 781-C terminus) was obtained from insect cells. Thecatalytic hydrolysis of ATP is measured using a radiometric filterbinding method. Incubation of kinase with ³³[P]ATP and substrate leadsto incorporation of ³³-[P] into the substrate which can then beseparated from the other reaction components by filtration. Assays wereperformed using 10 μM ATP and in the absence or presence of 1, 0.3, or0.1 μM compound. Activity was expressed as % of inhibition of control.

Example 612 Ambit KinomeScan Screening

This assay is an ATP-site dependent competition binding assay in whichhuman kinases of interest are fused to a proprietary tag (T7bacteriophage). The amount of kinase bound to an immobilized,active-site directed ligand is measured in the presence and absence ofthe test compound. Ambit's JAK assays use kinase domains and notfull-length proteins. The domain used for JAK1 binding is the pseudokinase domain while that for JAK3 binding is the catalytic domain (MazenW Karaman, Sanna Herrgard, Daniel K Treiber, et. al. A Quantitativeanalysis of kinase inhibitotr selectivity. Nature Biotechnology, 2008,Volume 26, No. 1, Page 127-132).

TABLE 6b Potency and Specificity of Kinase Inhibition (IC50 in nM)Compound Syk Jak 1 Jak 2 Jak 3 P142-76  

 4 No inhibition at 500 nM No inhibition at 500 nM No inhibition at 500nM Example 87  

 6 No inhibition at 300 nM No inhibition at 300 nM No inhibition at 300nM Example 596  

43 No inhibition at 300 nM No inhibition at 300 nM No inhibition at 300nM

Example 605 JAK3/STAT6 Cellular Assay

Stimulation of Ramos B cells by interleukin 4 (IL4) leads to signalingthrough JAK1/JAK3 resulting in phosphorylation of STAT6 (signaltransducers and activators of transcription). The effect of compounds oninhibition of JAK3 and/or JAK1 can be assessed by measuring the amountof phosphorylated STAT6. This is performed by Western blotting using aspecific phospho-STAT6 antibody.

Ramos B cells were suspended in 10 mM Hepes-buffered RPMI media (2×10⁷cells/ml). Cells (90 μl) were incubated with 10 μl 3.3 μg/ml interleukin4 (R & D Systems Inc, cat #204-IL; final concentration: 0.33 μg/ml).Incubations were for 10 min at 37° C. in the absence or presence of 2 μlcompound diluted in 30% DMSO. Reactions were terminated by the additionof an equal volume of 2× lysis buffer (100 mM TRIS-HCl pH 8.0, 2%Triton-X-100, 5 mM EDTA, 250 mM NaCl, 20% glycerol, 1.25 mM PMSF, 5 mMsodium orthovandate, 5 mM (3-glycerophosphate, mini complete EDTAprotease inhibitor cocktail (Sigma)).

Samples were incubated with 1 μl of the nuclease, benzonase (Novagen,cat #71205-3) for 1 hour, room temperature and then 50 μl 5× loadingbuffer (330 mM TRIS pH 6.8, 9.5 SDS, 34% glycerol, 0.01% bromophenolblue, 10% beta-mercaptoethanol) was added.

Cell lysates (15 μL) were subjected to SDS-PAGE (Novex 4-12%TRIS-glycine gels, Invitrogen) under reducing conditions, followed byelectroblot-transfer onto nitrocellulose membranes. Membranes were thenincubated in Zymed blocking buffer (Invitrogen) for 1 hr at roomtemperature (RT) then overnight at 4° C. with 1:500 antiphosphotyrosine-STAT6 (Cell Signaling Technology, cat #9364) primaryantibody in Zymed blocking buffer. Following 5×10 mM washes withTris-buffered saline, 0.25% NP40 (TBSN), blots were incubated for 1 hrat room temperature in the presence of 1:10,000 HRP-conjugated donkeyanti-rabbit secondary antibody (Amersham Biosciences, cat #NA934V) inZymed blocking buffer. After 4×10 min TBSN washes, blots were visualizedby ECL (Pierce Western Lightening, Perkin Elmer cat #NEL101). In orderto determine total (33 content, blots were stripped, washed 4× withTBSN, and re-probed with 1:2000 C3A antibody in block buffer overnightat 4° C. After 4×10 min TBSN washes, blots were incubated with 1:10,000goat anti-mouse secondary antibody in blocking buffer, washed 4 moretimes with TBSN and exposed to Western Lightening reagent. Levels ofstimulation over background and the extent of inhibition of compoundwere were determined by densitometry.

Example 606 Inhibition of JAK Kinase Activity Assay for Ramos B-cellLine Stimulated with IL-4

These examples illustrate methods for evaluating the in vitro and invivo human JAK kinase activities of the inventive compounds can bedetermined by various procedures known in the art, such as a test fortheir ability to inhibit the activity of human plasma JAK kinase. Thepotent affinities for human JAK kinase inhibition exhibited by theinventive compounds can be measured by an IC₅₀ value (in nM). The IC₅₀value is the concentration (in nM) of the compound required to provide50% inhibition of human JAK kinase activity. The smaller the IC₅₀ value,the more active (potent) is a compound for inhibiting JAK kinaseactivity.

An in vitro assay for detecting and measuring inhibition activityagainst JAK kinase is as follows:

The activity of the compounds for JAK kinases is confirmed in cellularassays designed to test for JAK inhibition. Briefly, JAK inhibition istested in human Ramos B-cells activated with cytokine Interleukin-4(IL-4). Twenty to 24 hours post stimulation, the cells are stained forupregulation of CD23 and analyzed by FACS. Stimulation of the B-cellswith IL-4 leads to the activation of the JAK/STAT pathway throughphosphorylation of the JAK kinase JAK1 and JAK3, which in turnphosphorylate and activate transcription factors STAT-5 and STAT-6. Thelow-affinity IgE receptor (CD23) is upregulated by activated STAT-5.

For the assay, human Ramos B-cells (ATCC, Catalog No. CRL-1596) arecultured in RPMI 1640 medium (Cellgro, Catalog No. 10-040-CM) containing10% fetal bovine serum (JRH, Catalog No. 12106-500M) according to thepropagation protocol supplied with the cells, and maintained at adensity of approximately 3.5×10⁵ cells/ml. The day before the assay, thecells are diluted to 3.5×10⁵ cells/ml to insure they are in thelogorithmic growth phase. The cells are spun down, and suspended in RPMI1640 medium (Cellgro, MediaTech, Inc., Herndon, Va., Cat No. 10-040-CM)containing 5-10% fetal bovine serum (FBS), heat inactivated (JRHBiosciences, Inc, Lenexa, Kans., Cat No. 12106-500M) according to ATCCpropagation protocol. Cells are maintained at a density of 3.5×10⁴⁻⁵cells/ml. The day before the experiment, Ramos B-cells are diluted to3.5×10⁵ cells/mL to ensure that they are in a logarithmic growth phaseand aliquots dispensed into a 96-well tissue culture plate. Cells areincubated with test compound (dissolved in DMSO) or DMSO (control) for 1hr at 37° C. and then stimulated with IL-4 (Pepotech, Catalog No.200-04) for 20-24 hours (final concentration is 50 Units/ml).

Cells are spun down and suspended in RPMI with 5% serum. 5×10⁴ cells areused per point in a 96-well tissue culture plate. Cells arepre-incubated with compound or DMSO (Sigma-Aldrich, St. Louis, Mo., CatNo. D2650) vehicle control for 1 hour in a 37° C. incubator. Cells arethen stimulated with IL-4 (Peprotech Inc., Rocky Hill, N.J., Cat No.200-04) for a final concentration of 50 units/mL for 20-24 hours. Cellsare then spun down and stained with anti-CD23-PE(BD Pharmingen, SanDiego, Calif., Cat No. 555711) and analyzed by FACS. Detection isperformed using a BD LSR I System Flow Cytometer, purchased from BectonDickinson Biosciences of San Jose, Calif.

Proliferation is measured using CellTiter-Glo® Luminescent CellViability Assay (Promega), which determines the number of viable cellsin culture based on quantitation of the ATP present, as an indicator ofmetabolically active cells. The substrate is thawed and allowed to cometo room temperature. After mixing the Cell Titer-Glo reagent and diluenttogether, 100 μL is added to each well. The plates are mixed on anorbital shaker for two minutes to induce lysis and incubated at roomtemperature for an additional ten minutes to allow the signal toequilibrate. Detection is performed using a Wallac Victor2 1420multilabel counter purchased from Perkin Elmer, Shelton, Conn.

On day two, A549 cells are pre-incubated with a 2,4-pyrimidinediaminetest compound or DMSO (control) (Sigma-Aldrich, St. Louis, Mo., CatalogNo. D2650) for 1 hour. The cells are then stimulated with IFNγ (75ng/mL) (Peprotech Inc., Rocky Hill, N.J., Cat. No. 300-O₂) and allowedto incubate for 24 hours. The final test compound dose range is 30 μM to14 nM in 2004 F12K media containing 5% FBS, 0.3% DMSO.

On day three, the cell media is removed and the cells are washed with200 μL PBS (phosphate buffered saline). Each well is trypsinized todissociate the cells, then neutralized by addition of 200 μL completeF12K media. Cells are pelleted and stained with an APC conjugated mouseanti-human ICAM-1 (CD54) (BD Pharmingen, San Diego, Calif., Catalog#559771) antibody for 20 minutes at 4° C. Cells are washed with ice coldFACS buffer (PBS+2% FBS) and surface ICAM-1 expression is analyzed byflow cytometry. Detection is performed using a BD LSR I System FlowCytometer, purchased from BD Biosciences of San Jose, Calif. Events aregated for live scatter and the geometric mean is calculated(Becton-Dickinson CellQuest software version 3.3, Franklin Lakes, N.J.).Geometric means are plotted against the compound concentration togenerate a dose response curve.

Example 607 Inhibition of Syk-mediated Signal Transduction Through the BCell Receptor in Non-Hodgkin's Lymphoma Cell Lines

Cells were pre-treated for 1 hour without or with compound (0.02 to 2uM) prior to stimulation of B cell receptor singling by incubation ofcells with 3 μg/ml anti-mu antibody for 10 minutes at 37° C. Ca²⁺ fluxwas measured using the Calcium 3 loading dye and the FlexStation(Molecular Device). B cell receptor signaling was assayed byintracellular phospho-Flow Cytometry, following protocols supplied by BDPharmingen (San Jose, Calif.). Syk activation was measured by inductionof BLNK tyrosine phosphorylation at amino acid position 84 (pBLNK Y84)and induction of ERK1/2 tyrosine phosphorylation at amino acid position204 (pERK Y204). Activation of the Src family member Lyn was measured byinduction of Syk tyrosine phosphorylation at amino acid position 352(pSyk Y352). Data are presented as μM IC₅₀s. Each compound effectivelyinhibited B cell receptor-induced Ca²⁺ fluxing and activation of Syk,but not the Src family member Lyn.

TABLE 7A Ramos Example 87  

Ca²⁺ 0.117 pBLNK Y84 0.5-0.75 pERK Y204 0.02-0.125 pSyk Y352 0.75-2   

TABLE 7B SUDHL-6 Example 87  

Ca²⁺ 0.111 pBLNK Y84 0.1-0.3 pERK Y204 0.1 pSyk Y352 >2

TABLE 7C Ramos Example 596  

Ca²⁺ 0.123 pBLNK Y84 0.5-0.75 pERK Y204 0.02-0.125 pSyk Y352 >2   

TABLE 7D SUDHL-6 Example 596  

Ca²⁺ 0.057 pBLNK Y84 0.1-0.3 pERK Y204 0.1  pSyk Y352 >2   

Example 608 Syk Inhibition Exerts an Anti-proliferative Effect onNon-Hodgkin's Lymphoma Cell Lines

Cells were incubated with increasing concentrations of each compound,then evaluated at 72 hours for extent of proliferation using the MTTassay (company, city, state) following the manufacturer suppliedprotocol. Data are presented as 1 μM IC50 values, representing the meanplus/minus standard deviation from 5 or 6 independent experiments. Eachcompound inhibited proliferation of SUDHL-4 and -6 cell lines, whichrely on Syk for survival and growth signals, in the low μM range. Toledocells which do not require Syk, was noticeably less sensitive to theanti-proliferative effects of Syk inhibition.

TABLE 8 SUDHL-4 SUDHL-6 Toledo Example 87  

1.8 ± 0.7 (5) 1.1 ± 0.4 (5) 9.3 ± 4.0 (5) Example 596  

5.4 ± 1.8 (5) 2.6 ± 1.4 (5) 38 ± 19 (5)

Example 609 Syk Inhibition Induces Apoptosis in Non-Hodgkin's LymphomaCell Lines

Data represent two independent experiments to evaluate the effect of Sykand Syk/JAK inhibition on survival of diffuse large non-Hodgkin'slymphoma B cell lines. SUDHL-4 and SUDHL-6 cells lines rely onSyk-mediated B cell receptor signaling for survival, while Toledo cellsdo not. Cells were incubated with compounds at the indicatedconcentrations and times; induction of apoptosis was measured by flowcytometry using the Caspase 3 Detection Kit (Sigma-Aldrich, Saint Luis,Mo.). Data are presented as the percent of total cells positive for theapoptosis marker, caspase 3. As expected, Syk inhibition resulted in theinduction of apoptosis in SUDHL-4 and -6 cell lines, but not the Toledocell line.

TABLE 9A Example 87  

24 h 48 h 72 h SUDHL-4  1 uM  8.7 10.7  9.9  3 uM 18.5 32.3 32.6 10 uM34.5 59.4 75.3 SUDHL-6  1 uM 11   20.2 20.4  3 uM 22.5 54.9 71   10 uM12.1 27.3 38.7 Toledo  1 uM  0.6  0.3  1.2  3 uM  0.8 0   1.8 10 uM 1  3.3  4.3

TABLE 9B Example 87  

24 h 48 h 72 h SUDHL-4  1 uM  6.6  7.2  7.6  3 uM 13.6 19.7 23.4 10 uM20.4 43.9 57.4 SUDHL-6  1 uM 12.9 21.9 15    3 uM 19.9 39.5 34.6 10 uM12.5 19.9 23.5 Toledo  1 uM 0   0.2  0.5  3 uM 0   0.1  0.2 10 uM 0  0.9  2.5

TABLE 9C Example 596  

24 h 48 h 72 h  1 uM  1.1  0.9  5.8  3 uM  4.9  3.5  5.2 10 uM 12.4 16.715.5 SUDHL-6  1 uM  8.3 11.6 10.6  3 uM 15.1 23.7 17.8 10 uM 17.8 37.332.4 Toledo  1 uM 0   0.1  0.2  3 uM 0  0   0.1 10 uM 0   0.2  0.7

Example 610 Inhibition of Mouse Primary B Cell Activation by SykInhibitors

Mouse primary splenocytes were pre-treated for 1 hour with increasingconcentrations of each compound (0.05-2 μM) prior to addition of controlor goat anti-mouse IgD serum. Anti-IgD induced B cell activation wasmeasured 16 hours later by flow cytometry, staining for the activationmarkers CD80/86 and CD69. Data represent IC₅₀ ranges for the inhibitionof B cell activation.

TABLE 10 CD69 CD80/86 Example 87  

<0.05  0.05-0.125 Example 596  

<0.05 0.125-0.25 

TABLE 11 CD69 CD80/86 Example 87  

<0.05 0.125-0.25 Example 596  

0.125- 0.25 0.125-0.25

Example 611 Mouse Model of Immune-mediated Thrombocytopenia

Immune-mediated thrombocytopenia is caused by antibodies directedagainst platelet surface glycoproteins, antibodies againstdrug-containing complexes on the platelet surface, or by antibody-coatedcells or immune complexes that interact with the platelet surface.Select compounds were evaluated for their ability to inhibit plateletclearance in a mouse model of antibody-mediated thrombocytopenia. Inthis model, a rapid clearance of circulating platelets (approximately50%) results from the intravenous administration of a rat anti-mouseGPIIb (clone MWReg30) antibody (BD Biosciences, Pharmingen). To evaluatecapacity for inhibition of platelet clearance, compounds were suspendedinto 0.5% methycellulose in water and administered via oral gavage (100ul/mouse) at a time prior to antibody injection when the compound wouldachieve maximum plasma concentration (typically 1-2 hours based onseparate pharmacokinetic experiments for individual compounds). At 4 and8 hours after injection of antibody, terminal blood samples wereobtained from groups of vehicle and test article treated mice (n=5-10mice/group) via cardiac puncture. Blood was anticoagulated usingtrisodium citrate or EDTA. Whole blood samples were measured forplatelet counts on a hematology analyzer (Hemavet, Drew Scientific).Remaining blood was processed for plasma and compound concentrationsmeasured by mass spectrometry.

Platelet clearance was determined by measuring the difference inplatelet number between the average non-antibody treatment group andanimals administered the rat anti-mouse GPIIb antibody. Inhibition ofplatelet clearance was determined by comparing the difference betweenplatelet clearance of vehicle and compound treated animals.

TABLE 12 AVERAGE AVERAGE INHIBITION INHIBITION AVERAGE OF (%) OF PEAKPLASMA PLATELET PLATELET CONCENTRATION CLEARANCE CLEARANCE COMPOUND DOSE(μM) (4 HRS) (8 HRS) Example 87  

  30 MG/KG/DAY 27%* 12% Example 87  

  10 MG/KG/DAY 7% 13% *DENOTES STATISTICALLY SIGNIFICANT REDUCTION INCLINICAL INFLAMMATION SCORE COMPARED TO VEHICLE CONTROL BY 2-TAILED,UNPAIRED, STUDENTS T-TEST

Average Peak Plasma Average Inhibition (%) of Compound doseConcentration (μM) inflammation score Example 596  

  100 mg/kg/day 10.2 μM 88%* EXAMPLE 596  

  30 mg/kg/day  4.4 μM 27%* *denotes statistically significant reductionin clinical inflammation score compared to vehicle control by 2-tailed,unpaired, Students t-Test

Example 612 Mouse Model of Collagen Antibody Induced Arthritis

The inhibitory activity of select compounds was investigated in a mousemodel of collagen antibody induced arthritis (CAIA). Collagen inducedarthritis is mediated by autoantibodies to type II collagen andcomplement, thus arthritis can be induced by administration ofpolyclonal antibodies or a mixture of monoclonal antibodies to type IIcollagen. The CAIA model (Chondrex, Inc., Redmond, Wash.) uses a mixtureof 4 clones which recognize individual epitopes clustered within an 83amino acid peptide fragment of type II collagen. These epitopes sharecommon amino acid sequences with many different species of type IIcollagen including chicken, mouse, rat, bovine, porcine, monkey andhuman. The model utilizes a monoclonal antibody cocktail followed bybacterial lipopolysaccharide (LPS) to induce a severe and consistentarthritis in mice within 7 days. This model was developed based on thehypothesis that bacterial toxins absorbed through the gastrointestinaltract play a synergistic and pathologic role with autoantibodies to typeII collagen in triggering arthritis in patients with RheumatoidArthritis.

For these experiments, the monoclonal antibody cocktail (Lot # OC-708)was injected intravenously via tail vein at a dose of 4 mg/mouse (40mg/ml) on day 0 followed by intraperitoneal injection of LPS dilutedinto normal saline at a dose of 25 ug/mouse in 8 week old, female Balb/Cmice (Charles River, Inc.). Dosing of test articles was started justbefore or after the IV injection of antibody cocktail. Compounds weresuspended into 0.5% methylcellulose in water and administered via oralgavage (100 ul/mouse) daily for the duration of the 7-10 day study.Clinical inflammation scores were obtained daily. Inhibition of clinicalinflammation scores was determined based on the difference betweenvehicle and test article treated mice at the end of the experiment.Plasma concentrations represent peak concentration at 1 hour post lastdose on the day of study termination.

TABLE 13 AVERAGE PEAK AVERAGE PLASMA INHIBITION (%) OF CONCENTRATIONINFLAMMATION COMPOUND DOSE (μM) SCORE Example 87  

  30 mg/kg/day 7.8 μM 44%* *denotes statistically significant reductionin clinical inflammation score compared to vehicle control by 2-tailed,unpaired, Students t-Test

Example 613 Inhibition of IL-4 Induced JAK1/3 to Stat-6 Phosphorylationin Ramos B Cells

Ramos B cells were pre-treated for 1 hour with increasing concentrationsof compound, as indicated prior to addition of IL-4. Cells wereincubated with IL-4 for 10 minutes, and then subjected to intracellularflow cytometry to measure the percent inhibition of IL-4 induced Stat-6.

Example 614 Inhibition of IL-4 Induced JAK1/3 to Stat-6 Phosphorylationin Ramos B Cells

Ramos B cells were pre-treated for 1 hour with increasing concentrationsof compound, as indicated prior to addition of IL-4. Cells wereincubated with IL-4 for 10 minutes, and then subjected to intracellularflow cytometry to measure the percent inhibition of IL-4 induced Stat-6.

Example 615 Primary Human T-Cell Proliferation Assay Stimulated withIL-2

Primary human T-cells derived from peripheral blood and pre-activatedthrough stimulation of the T-cell receptor and CD28 proliferate in vitroin response to the cytokine Interleukin-2 (IL-2). This proliferativeresponse is dependent on the activation of JAK-1 and JAK-3 tyrosinekinases, which phosphorylate and activate the transcription factorStat-5.

Human primary T cells are prepared as follows. Whole blood is obtainedfrom a healthy volunteer, mixed 1:1 with PBS, layered on to FicollHypaque (Amersham Pharmacia Biotech, Piscataway, N.J., Catalog#17-1440-03) in 2:1 blood/PBS:ficoll ratio and centrifuged for 30 min at4° C. at 1750 rpm. The lymphocytes at the serum: ficoll interface arerecovered and washed twice with 5 volumes of PBS. The cells areresuspended in Yssel's medium (Gemini Bio-products, Woodland, Calif.,Catalog #400-103) containing 40 U/mL recombinant IL2 (R and D Systems,Minneapolis, Minn., Catalog #202-IL (20 μg)) and seeded into a flaskpre-coated with 1 μg/mL anti-CD3 (BD Pharmingen, San Diego, Calif.,Catalog #555336) and 5 μg/mL anti-CD28 (Immunotech, Beckman Coulter ofBrea Calif., Catalog #IM1376). The primary T-cells are stimulated for 3to 4 days, then transferred to a fresh flask and maintained in RPMI with10% FBS and 40 U/mL IL-2.

Primary T-cells are washed twice with PBS to remove the IL-2 andresuspended in Yssel's medium at 2×10⁶ cells/mL. 50 μL of cellsuspension containing 80 U/mL IL-2 is added to each well of a flatbottom 96 well black plate. For the unstimulated control, IL-2 isomitted from the last column on the plate. Compounds are seriallydiluted in dimethyl sulfoxide (DMSO, 99.7% pure, cell culture tested,Sigma-Aldrich, St. Louis, Mo., Catalog No. D2650) from 5 mM in 3-folddilutions and then diluted 1:250 in Yssel's medium. 50 μL of 2× compoundis added per well in duplicate and the cells are allowed to proliferatefor 72 hours at 37° C.

Proliferation is measured using CellTiter-Glo® Luminescent CellViability Assay (Promega), which determines the number of viable cellsin culture based on quantitation of the ATP present, as an indicator ofmetabolically active cells. The substrate is thawed and allowed to cometo room temperature. After mixing the Cell Titer-Glo reagent and diluenttogether, 100 μL is added to each well. The plates are mixed on anorbital shaker for two minutes to induce lysis and incubated at roomtemperature for an additional ten minutes to allow the signal toequilibrate. Detection is performed using a Wallac Victor2 1420multilabel counter purchased from Perkin Elmer, Shelton, Conn.

Example 616 A549 Epithelial Line Stimulated with IFNγ

A549 lung epithelial cells up-regulate ICAM-1 (CD54) surface expressionin response to a variety of different stimuli. Therefore, using ICAM-1expression as readout, compound effects on different signaling pathwayscan be assessed in the same cell type. IFNγ up-regulates ICAM-1 throughactivation of the JAK/Stat pathway. In this example, the up-regulationof ICAM-1 by IFNγ is assessed.

The A549 lung epithelial carcinoma cell line originated from theAmerican Type Culture Collection. Routine culturing is with F12K media(Mediatech Inc., Lenexa, Kans., Cat. No. 10-025-CV) with 10% fetalbovine serum, 100 I.U. penicillin and 100 ng/mL streptomycin (completeF12k media). Cells are incubated in a humidified atmosphere of 5% CO₂ at37° C. Prior to use in the assay, A549 cells are washed with PBS andtrypsinized (Mediatech Inc., Cat. No. 25-052-CI) to lift the cells. Thetrypsin cell suspension is neutralized with complete F12K media andcentrifuged to pellet the cells. The cell pellet is resuspended incomplete F12K media at a concentration of 2.0×10⁵/mL. Cells are seededat 20,000 per well, 100 μL total volume, in a flat bottom tissue cultureplate and allowed to adhere overnight.

On day two, A549 cells are pre-incubated with a 2,4-pyrimidinediaminetest compound or DMSO (control) (Sigma-Aldrich, St. Louis, Mo., CatalogNo. D2650) for 1 hour. The cells are then stimulated with IFNγ (75ng/mL) (Peprotech Inc., Rocky Hill, N.J., Cat. No. 300-O₂) and allowedto incubate for 24 hours. The final test compound dose range is 30 μM to14 nM in 200 μL F12K media containing 5% FBS, 0.3% DMSO.

On day three, the cell media is removed and the cells are washed with200 μl PBS (phosphate buffered saline). Each well is trypsinized todissociate the cells, then neutralized by addition of 200 μL completeF12K media. Cells are pelleted and stained with an APC conjugated mouseanti-human ICAM-1 (CD54) (BD Pharmingen, San Diego, Calif., Catalog#559771) antibody for 20 minutes at 4° C. Cells are washed with ice coldFACS buffer (PBS+2% FBS) and surface ICAM-1 expression is analyzed byflow cytometry. Detection is performed using a BD LSR I System FlowCytometer, purchased from BD Biosciences of San Jose, Calif. Events aregated for live scatter and the geometric mean is calculated(Becton-Dickinson CellQuest software version 3.3, Franklin Lakes, N.J.).Geometric means are plotted against the compound concentration togenerate a dose response curve.

Example 617 U937 IFNγICAM1 FACS Assay

U937 human monocytic cells up-regulate ICAM-1 (CD54) surface expressionin response to a variety of different stimuli. Therefore, using ICAM-1expression as readout, compound effects on different signaling pathwayscan be assessed in the same cell type. IFNγ up-regulates ICAM-1 throughactivation of the JAK/Stat pathway. In this example, the up-regulationof ICAM-1 by IFNγ is assessed.

The U937 human monocytic cell line is obtained from ATCC of Rockville,Md., catalog number CRL-1593.2, and cultured in RPMI-1640 mediumcontaining 10% (v/v) FCS. U937 cells are grown in 10% RPMI. The cellsare then plated at a concentration of 100,000 cells per 160 μL in 96well flat bottom plates. The test compounds are then diluted as follows:10 mM test compound is diluted 1:5 in DMSO (3 μL 10 mM test compound in12 μL DMSO), followed by a 1:3 serial dilution of test compound in DMSO(6 μL test compound serially diluted into 12 μL DMSO to give 3-folddilutions). Then 4 μL of test compound is transferred to 76 μL of 10%RPMI resulting in a 10× solution (100 μM test compound, 5% DMSO). Forcontrol wells, 4 μL of DMSO is diluted into 76 μL 10% RPMI.

The assay is performed in duplicate with 8 points (8 3-fold dilutionconcentrations from 10 μL) and with 4 wells of DMSO only (control wells)under stimulated conditions and 4 wells of DMSO only under unstimulatedconditions.

The diluted compound plate is mixed 2× using a multimek (Beckman Coulterof Brea, Calif.) and then 20 μL of the diluted compounds is transferredto the 96 well plate containing 160 μL of cells, which are then mixedagain twice at low speeds. The cells and compounds are thenpre-incubated for 30 minutes at 37° C. with 5% CO₂.

The 10× stimulation mix is made by preparing a 100 ng/mL solution ofhuman IFNγ in 10% RPMI. The cells and compound are then stimulated with20 μL of IFNγ stimulation mix to give a final concentration of 10 ng/mLIFNγ, 10 μM test compound, and 0.5% DMSO. The cells are kept underconditions for stimulation for 18-24 hours at 37° C. with 5% CO₂.

The cells are transferred to a 96 well round bottom plate for stainingand then kept on ice for the duration of the staining procedure. Cellsare spun down at 1000 rpm for 5 minutes at 4° C., following which thesupernatant is removed. Following removal of the supernatant, 1 μL APCconjugated mouse anti-human ICAM-1 antibody is added per 100 μL FACSbuffer. The cells are then incubated on ice in the dark for 30 minutes.Following incubation, 150 μL of FACS buffer is added and the cells arecentrifuged at 1000 rpm for 5 minutes at 4° C., following which thesupernatant is removed. After removal of the supernatant, 200 μL of FACSbuffer is added and the cells are resuspended. After suspension, thecells are centrifuged at 1000 rpm for 5 min at 4° C. Supernatant is thenremoved prior to resuspension of the cells in 150 μL FACS buffer.

Detection is performed using a BD LSR I System Flow Cytometer, purchasedfrom BD Biosciences of San Jose, Calif. The live cells are gated forlive scatter and the geometric mean of ICAM-APC is measured(Becton-Dickinson CellQuest software version 3.3, Franklin Lakes, N.J.).Both % live cells and ICAM-1 expression is analyzed. The assays for thetest compounds is carried out in parallel with a control compound ofknown activity. The EC₅₀ for the control compound is typically 40-100nM.

Example 618 Analysis of B Cell Signaling

The human non-Hodgkin's lymphoma B cell lines SUDHL-4 (#ACC 495),SUDHL-6 (#ACC572), and Karpas-422 (#ACC32) were obtained from DSMZ(Braunschweig, Germany); Toledo (#CRL-2631) and Ramos (#CRL-1596) wereobtained from the American Type Culture Collection (ATCC; Manassas,Va.). All cells were maintained in RPMI media (Invitrogen, Carlsbad,Calif.) supplemented with 10% fetal calf serum (ATCC) andpenicillin/streptomycin (Invitrogen), and maintained in a 37° C.humidified tissue culture incubator. Antibodies used in these studiesinclude polyclonal goat F(ab)′2 anti-human IgG (H+ L) and anti-human IgM(BioSource, Camarillo, Calif.); rabbit anti-human Syk, rabbit anti-humanphospho-Syk (Y525/526), rabbit anti-human phospho-Syk (Y352), anti-humanBLNK, anti-human phospho-BLNK (Y84) were obtained from Cell SignalingTechnologies, Inc. (Danvers, Mass.). The following antibodies wereobtained from Becton Dickenson (San Jose, Calif.) for phospho-flowcytometry: Alexa fluor 488-conjugated mouse anti-human phospho-STAT6(Y641), Phycoerythrin (PE)-conjugated mouse anti-human phospho-Zap70(Y319)/Syk(Y352), and Fluorescein isothiocyanate (FITC)-conjugated mouseanti-human phospho-ERK1/2 (T202/Y204).

Phospho-flow cytometry was performed essentially as described elsewhere(Irish, Czerwinski et al. Blood 108(9): 3135-42 (2006). 0.5×10⁶ cells ingrowth media were stimulated with 1 μg/ml anti-μ or anti-γ antibody for10 minutes. Induced signaling was terminated immediately following theindicated time by the addition of paraformaldehyde (Electron MicroscopySciences, Hatfield, Pa.) to a final concentration of 1%. Cells wereincubated with paraformaldehyde for 5 minutes at room temperature,washed once with phosphate buffered saline (PBS), then resuspended andincubated overnight at 4° C. in pre-chilled methanol (−80° C.) (company,address). Fixed and permeablized cells were subsequently washed once inPBS, a second time in PBS containing 1% bovine serum albumin (BSA)(Sigma-Aldrich, St. Louis, Mo.), and then stained with the indicatedantibodies diluted 1:20 in PBS+1% BSA. After 30 minutes, cells werewashed once in PBS and subjected to flow cytometry using the FACSCalibur (Becton Dickenson). For Western blot analyses, 106 cells werestimulated for 30 minutes with 2 μg/ml of the indicated BCR-specificantibodies. Signaling was terminated by resuspending the cells in lysisbuffer and incubated on ice for 1 hour. Cell debris were removed bycentrifugation, and the cleared protein lysates were resolved by 10%SDS-PAGE and probed with the indicated antibodies followingrecommendations made by the manufacturers. Where indicated, cells werepre-treated for 1 hour at 37° C. with Syk inhibitors or vehicle control(0.5% DMSO) at several concentrations prior to stimulation with anti-BCRantibody.

Example 619 Selective Inhibition of Syk Activity

Compounds were tested for their ability to inhibit purified Syk. example596 and example 87 (two compounds from a Syk-specific series as shown inTable 1b) and P420-89 (from a series with dual Syk and JAK inhibitoryactivities) were found to suppress Syk kinase activity with IC50s of 43nM, 6 nM, and 31 nM, respectively. The selectivity of these compoundsfor Syk was determined by screening each against a panel of 270independent purified kinases at 300 nM (Millipore). The percentinhibition relative to vehicle control was calculated, and the numberswere converted into a heat-map; no inhibition is represented as green,increasing blending with red indicates increasing percent inhibitionwith yellow representing 50% inhibition and red representing 100%inhibition (FIG. 8). As depicted in FIG. 8A, example 596 and example 87were highly Syk specific (first and second rows, respectively) whereasP420-89 inhibited multiple kinases (third row). The subset of kinasesthat were inhibited by >80% by any of the three compounds are shown inFIG. 8B. example 596 inhibited Syk and MLK-1 (first row). At 300 nMexample 87 inhibited 10 different kinases (second row). When re-testedat 50 nM (approximately 10× above its Syk 1050 value of 6 nM), however,Syk was the only kinase that remained inhibited (third row). P420-89inhibited Syk, JAK2 and JAK3, along with several other kinases (fourthrow).

Employing the Milipore panel of purified kinases EXAMPLE 87 (IC₅₀=1 nM)inhibited 98% of purified Syk kinase activity at 50 nM. IC50 values weredetermined for those kinases that were inhibited by >80% at 300 nM inthe Millipore kinase panel.

Kinase IC50 (nM) Syk(h) 1 MLK1 60 Fgr(h) 81 Yes(h) 123 Flt3(h) 139 PAK5166 Lyn(h) 199 cSRC(h) 244 Lck(h) 300

By contrast, multi-kinase inhibitor P420-89 is more akin to Rigel'sR788. At 300 nM, P420-89 inhibited Syk by 88%, along with >80%inhibition of 32 additional kinases. Among these were JAK 2 and 3 (93%and 85% inhibited, respectively), Flt-3 (83-92% inhibited), and cKit(95-97% inhibited), all targets for therapeutic manipulation oflymphocyte function.

Example 620 Calcium Flux Assay and Selective Inhibition of Syk inNon-Hodgkin's Lymphoma B Cell Lines

Ramos cells were cultured (maintaining approximately 0.5×10⁶ cells/ml)in growth medium 3 to 4 days ahead of experiments. Cells were harvestedand re-suspended in fresh medium at 8×10⁶ cells/ml before dye-loading.An equal volume of Calcium 3 loading dye (Molecular Device, Sunneyvale,Calif.) was added to the cell suspensions. Loaded cells were dispensedin a 96 well plate and incubated for 20 minutes. Syk inhibitors werethen added to the loaded cells and incubated for another 30 minutes. Bcells were stimulated with 5 μg/ml anti-μ antibody. Changes inintracellular Ca2⁺ concentration was measured using the FlexSTATion(Molecular Devices, Sunnyvale, Calif.).

The electivity and potency of Syk inhibition in B cells was initiallyinterrogated by Western blot, measuring BCR-mediated induction of pSykY525/526 and pBLNK Y84, both measures of Syk kinase activity, and theinduction of pSyk Y352, a measure of Src kinase activity. SUDHL-6 Bcells were stimulated with anti-BCR specific antibody for 30 minutes inthe presence or absence of each Syk inhibitor or vehicle control.Treatment with 0.16 or 1 M of each compound reduced BCR-induced Sykautophorphorylation (Y525/526) by roughly 40% and 60%, respectively, asestimated by densitometry (data not shown). An expanded range ofconcentrations was used to further evaluate the effect of thesecompounds on BCR induced Syk and Src kinase activity. As shown in FIG.9, A-C, each compound inhibited Syk activity (pBLNK Y84) with IC50values ranging from 0.16 to 1 M, while no effect on Src activity (pSykY352) was observed as high as 2.5 M.

The ability of each compound to suppress signaling events more distal tothe BCR was also measured. Cells were again stimulated by anti-BCRantibody in the presence or absence of various concentrations of eachSyk inhibitor. The induction of pSyk Y352 was measured as a specificitycontrol, while that of pERK1/2/Y204 was used as a measure of more distalSyk-dependent signaling (Jiang, Craxton et al. J Exp Med 188(7):1297-306 (1998). FIG. 12C shows representative FACS plots depicting theeffect of the most specific and potent Syk inhibitor of the three,example 87, on BCR signaling. At 125 nM, ERK1/2 activation wascompletely suppressed, whereas the stimulated cells still stainedpositive for Syk Y352. This experiment was repeated, in which the effectof all three compounds on Src and Syk activity were determined (FIG.10A). Concentrations of less than 125 nM were sufficient to suppress BCRinduced Syk signaling to ERK1/2. By contrast, much higher concentrationswere required to cause a modest suppression of Src activity; an effecton Src that was not observed by Western blot (FIG. 9, A-C). None ofthese Syk inhibitors suppressed PMA-induced ERK1/2 tyrosinephosphorylation, demonstrating these compounds do not inhibit signalingevents down-stream of PKC.

Whereas example 596 and example 87 specifically inhibited Syk inpurified and cellular assays, P420-89 additionally demonstrated activityagainst purified JAK kinases. These compounds were tested for inhibitionof IL-4 signaling to STAT-6 via JAK1/3 in B cells, a signaling pathwaythat does not require Syk. The Syk specific compounds did not suppressIL4 signaling at concentrations as high as 2 μM. Conversely, P420-89 didsuppress IL4 signaling, with an IC50 around 125 nM (FIG. 10B).

This shows that selective inhibition of Syk suppressed BCR-induced Ca2+flux in B cells with IC50 values around 100 nM. This suggests that byinhibiting Syk, these compounds suppress the signaling pathway, blockingthe cellular response.

Selective inhibition of Syk is sufficient to suppress BCR signalingwithout affecting Src (FIGS. 9, 10 and 12) or JAK (FIG. 10B). Example 87suppressed proliferation of NHL cell lines with equal potency to themulti-kinase inhibitor, P420-89 (Table, Figure 16). Additionally,example 87 and P420-89 equally induced apoptosis in these cells (FIG.11B). This data demonstrates the role of Syk signaling in the survivalof NHL cell lines, and demonstrates that inhibition of kinases otherthan Syk is not required to achieve this effect.

Example 621 Caspase 3 and Proliferation Assays: Syk Inhibition DisruptsProliferation and Survival of Non-Hodgkin's Lymphoma B Cell Lines.

Induction of apoptosis was measured using the PE-conjugated monoclonalactive caspase-3 antibody apoptosis kit (Becton Dickenson) following thesupplied protocol. Cells were suspended in growth media (0.5×10⁶cells/ml) and treated with the indicated concentrations of each Sykinhibitor or vehicle control for 24, 48, or 72 hours prior to FACSanalysis. The MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide, a tetrazole) assay (company name) was used as a measure of cellviability and growth, following protocols supplied by the manufacturer.Cells were treated with the indicated concentrations of each Sykinhibitor or vehicle control for 72 hours.

SUDHL-4 and SUDHL-6 cells were previously classified as “BCR-type”(Monti, Savage et al. Blood 105(5): 1851-61 (2005); Polo, Juszczynski etal. Proc Natl Acad Sci U S A 104(9): 3207-12 (2007) and sensitive to Sykinhibition by R406 (Chen, Monti et al. 2008). The Toledo and Karpas-422cell lines that lack BCR and BLNK expression, respectively (Gabay,Ben-Bassat et al. Eur J Haematol 63(3): 180-91 (1999); Sprangers,Feldhahn et al. Oncogene 251(36): 5056-62 (2006), having thereforeadapted to survive independent of BCR signals, were insensitive to R406(Chen, Monti et al. 2008). The proliferation of these cell lines whencultured in the presence or absence of various concentrations of eachSyk inhibitor for 72 hours was tested. As presented in the Table in FIG.16, each compound suppressed proliferation of the Syk-dependent SUDHLcell lines with 1050 values in the low μM range. Toledo cells requiredmuch higher concentrations to affect proliferation. Dual suppression ofSyk and JAK kinases by P420-89 did not appear to have a greateranti-proliferative effect relative to Syk inhibition alone.

Selective inhibition of Syk was sufficient to induce apoptosis in“BCR-type” NHL cell lines. Cells were incubated with 1 or 3 M of the Sykspecific inhibitor example 87 for 72 h. As demonstrated in FIG. 11A,SUDHL-4 and -6 cells each underwent apoptosis, whereas the Toledo andKarpas-422 cells did not (FIG. 11A). In replicate experiments, thespecific inhibition of Syk by example 596 and example 87 inducedapoptosis only in the SUDHL and Ramos cell lines. By comparison,P420-89, which potently inhibits Syk and JAK kinases, induced apoptosisin all the “BCR-type” cell lines, as well as in Karpas-422 and JJN-3, amultiple myeloma cell line that lacks BCR, and BLNK expression(Sprangers, Feldhahn et al. Oncogene 25(36): 5056-62 (2006). The Toledocells remained insensitive to all three compounds (FIG. 11B). In aseparate experiment, the SUDHL-6 and Toledo cells were found to beequally sensitive to induction of apoptosis by 72 h treatment with 1 MPMA. These data demonstrate the specific requirement of Syk in thesurvival of certain NHL cell lines.

Example 622 Xenograft Studies and Tumor and Plasma ConcentrationAnalysis

Syk Inhibition Protects Against Tumor Formation in a Xenograft MouseModel. Mice were received (company) and acclimated in-house at leastthree days prior to use. Ramos cells (3×106) were injectedsubcutaneously into the hind flank area of conscious mice using a 27gauge needle in an injection volume of less than 0.5 ml. Followinginjection, mice were randomized into treatment groups (n=15) and dosedtwice daily by oral gavage with vehicle or 10, 15, or 20 mg/kg of theSyk inhibitor example 87. Body weights were obtained at least once perweek and caliper measurements of tumors were determined twice per weekbeginning when palpable tumorrs were formed until the end of the study.Tumor volume was assessed by caliper measurement using a formula[maximum length×width×height×π/6]. Twice daily dosing of vehicle orexample 87 continued until the vehicle or any treatment group exhibitedtumors that exceeded 1.5 grams in size. At the time of termination (5weeks post Ramos innoculation) the mice were anesthetized with aketamine cocktail. A blood sample was obtained for CBC and plasmaconcentration determination via cardiac puncture and the mice wereeuthanized via cervical dislocation. Tumors were then be excised andweighed. One half of the tumor was snap frozen in liquid nitrogen fordetermination of concentration of example 87 in the tumor tissue and theother half was placed in 10% buffered formalin for histologicalinvestigation.

The effect of Syk inhibition on Ramos tumor formation in a xenograftmouse model was assessed. Mice were dosed twice daily with 10, 15, or 20mg/kg example 87 or vehicle control beginning the day of tumor cellinoculation. Caliper measurements were initiated when tumors began toform, approximately three weeks post-tumor inoculation, and repeatedevery third day until termination of the study. The study was terminatedwhen tumor weights began reaching approximately 1.5 mg, at which timetumors were excised and weighed. Tumor and plasma samples were subjectedto pharmacokinetic analysis.

Each tumor sample was homogenized in 3 ml of saline per gram of tumorusing the Kontes® Microtube Pellet Pestle® Rods and Motor (Kimble Chase,Vineland, N.J.). Plasma and tumor samples were analyzed for example 87concentration using a liquid chromatography tandem mass spectrometer(LC/MS/MS). In brief, plasma and tumor samples were processed in a96-well Captiva™ filter plate (0.2 μm, Varian, Inc., Palo Alto, Calif.).Aliquots of plasma and homogenized tumor samples were precipitated withacetonitrile containing 200 ng/mL of:

the internal standard. The mixture was vortexed and refrigerated at 4°C. for 30 minutes to allow complete protein precipitation. The mixturewas filtered into a 96-well collection plate. The filtrate was injectedonto a Sciex API3000 LC/MS/MS equipped with a turbo-ion spray source.example 87 and Compound A were separated on a Phenomenex Luna 5μ HILICcolumn (4.6×100 mm, 5 mm; Phenomenex, Torrance, Calif.). A mobile phasegradient mixture of 10% mobile phase A (0.1% formic acid in water) and90% mobile phase B (0.1% formic acid in 90% acetonitrile, 10% water) to65% mobile phase B was programmed over 1.1 minutes followed by agradient of mobile phase B from 65% to 90% over 0.01 minutes. The peakareas of the m/z 394/360 product ion of example 87 were measured againstthose of the m/z 357/295 product ion of Compound A (internal standard)in positive ion mode. The analytical range was 2 to 5000 ng/ml.

Pharmacokinetic analysis revealed that at steady-state, tumorconcentrations of example 87 followed the concentration-time profilesseen with plasma in the 10, 15, and 20 mg/kg dose groups. Nonlinearincreases in C_(max), AUC (0-8), and tumor C_(min) were observed as thedose was increased, but a dose-proportional increase in plasma C_(min)was noted. Mean C_(max) and AUC (0-8) in plasma was at least 2-foldgreater than that in tumor for all doses examined; however, mean nadirconcentrations (C_(min)) were higher in tumor than in plasma (Table14A), indicating accumulation of example 87 in the tumor compartment.

Table 14A Tmax Cmin Cmax AUC (0-8) Dosing regiment (hr) (ng/mL) (ng/mL)(ng * hr/mL) Determined from plasma 10 mg/kg BID 1.50 17.6 179 738 15mg/kg BID 1.50 26.6 343 1671 20 mg/kg BID 4.00 39.5 570 3191 Determinedfrom tumor 10 mg/kg BID 8.00 24.5 55.2 353 15 mg/kg BID* 4.00 67.8 163475 20 mg/kg BID 4.00 125 252 1453 Table 14B Dosing tumor/plasma ratioregimen AUC based Cmax based Cmin based 10 mg/kg BID 0.478 0.308 1.39 15mg/kg BID* 0.284 0.475 2.55 20 mg/kg BID 0.455 0.442 3.15 Note: Nadir(0), 1.5, 4, and 8 h samples were taken on the da of harvest followingthe AM dose. The second dose was not administered on the day of harvest;therefore, pharmacokinetic values above were determined after a singleAM dose at steady-state. *Only one tumor sample was available for the 8h time-point and may have been an outlier (tumor concentrations at 8 h -608 ng/ml); therefore, pharmacokinetic parameters were determinedbetween 0 to 4 h for the 15 mg/kg BID EXAMPLE 87 dose group. As aresult, AUC (0-8) and AUC based tumor/plamsa ration for this dose groupmay be underestimated.The difference between plasma and tumor C_(min) became more prominent asthe dose was increased, as indicated by the increase in tumor/plasmaratios determined from C_(min) (Table 14B). Tumor/plasma ratiosdetermined from C_(max) and AUC (0-8) were similar across the variousdose groups. Tumor concentrations were sustained above 60, 170, and 640nM over the entire dosing interval at steady-state for example 87 at 10,15, and 20 mg/kg, respectively.

Mice dosed with all three concentrations of example 87 were protectedfrom Ramos tumor growth in vivo. This was first evident from calipermeasurements (data not shown), which revealed a reduced rate of tumorgrowth in the presence of the Syk inhibitor. Upon study completion, micewere euthanized and tumors excised and weighed; data presented in FIG.17. Consistent with caliper measurements, a statistically significantreduction in average tumor weight was achieved in all dosing groups,relative to vehicle control. These data reveal that sub-micromolarconcentrations of example 87 can prevent tumor formation by anaggressive NHL cell line in mice.

Mice dosed with the Syk inhibitor did not present with reduced numbersin any subset of white blood cells. In fact, the only effect observedwas an increase in the number of lymphocytes in mice treated with15mg/kg example 87, which was not repeated in mice dosed with 10 or20mg/kg (FIG. 17). The relative percent of each cell subtype analyzedwas also unaffected by the Syk inhibitor (data not shown). On average,mice treated with vehicle control had a 9.45% increase in body weight.Mice treated with 10, 15, and 20mg/kg example 87, on the other hand, hadon average 0.27% increase, 1.67% decrease, and 2.27% decrease in bodyweight, respectively, over the course of the study. There was norelationship, however, between % change in body weight and tumor growth(R2=27). These data suggest that the inhibition of tumor growth wasindeed mediated by suppression of Syk activity.

The Syk-specific inhibitor example 87 was also tested for activity in aRamos tumor mouse xenograft model. At all the concentrations tested,statistically significant reductions in tumor growth were observed inmice dosed BID with example 87. The lowest concentration tested was 10mg/kg, achieving tumor concentrations ranging from 64 to 140 nM over thecourse of the day. Suppression of tumor growth at these concentrationsin vivo is consistent with concentrations of <125 nM found to suppressBCR-induced Ca2+ flux and distal BCR signaling to pERK Y204 (FIGS. 10and 12). The selective pharmacological inhibition of Syk results ineffects on the proliferations and survival of NHL cell lines. These datasuggest that the selective targeting of Syk may similarly have clinicalbenefit in a variety of B-cell proliferative disorders.

As detailed herein, Syk has been implicated experimentally in B celldevelopment, proliferation, and survival. Moreover, Syk is implicated asan oncogene. Expression of constitutively active Syk in adoptivelytransferred bone marrow cells induces leukemia in mice, andover-activity of Syk is associated with a variety of lymphomas in humansGiven the role of Syk in B cell biology, its selective inhibition may besufficient to provide clinical benefit in B cell proliferativedisorders, while reducing toxicities that may arise due to suppressionof other off-target kinases.

The present invention provides a number of embodiments. It is apparentthat the examples may be altered to provide other embodiments of thisinvention. Therefore, it will be appreciated that the scope of thisinvention is to be defined by the appended claims rather than by thespecific embodiments, which have been represented by way of example.

All of the above U.S. patents, U.S. patent application publications,U.S. patent applications, foreign patents, foreign patent applicationsand non-patent publications referred to in this specification and/orlisted in the Application Data Sheet, are incorporated herein byreference, in their entirety. From the foregoing it will be appreciatedthat, although specific embodiments of the invention have been describedherein for purposes of illustration, various modifications may be madewithout deviating from the spirit and scope of the invention.Accordingly, the invention is not limited except as by the appendedclaims.

1. A compound having the formula:

or a tautomer or a pharmaceutically acceptable salt thereof.
 2. Acomposition comprising a compound of claim 1 or a tautomer or apharmaceutically acceptable salt thereof in combination with apharmaceutically acceptable carrier or diluent.